155 results on '"Horst Kleinkauf"'
Search Results
2. The Roots of Modern Biochemistry
- Author
-
Horst Kleinkauf, Hans von Döhren, Lothar Jaenicke, Horst Kleinkauf, Hans von Döhren, Lothar Jaenicke
- Published
- 2011
3. Catabolism of Thyroliberin by Rat Adenohypophyseal Tissue Extract
- Author
-
Karl Bauer and Horst Kleinkauf
- Subjects
Male ,Stereochemistry ,Pyroglutamyl-Peptidase I ,In Vitro Techniques ,Aminopeptidases ,Biochemistry ,Aminopeptidase ,Enzyme catalysis ,Pyroglutamate aminopeptidase ,chemistry.chemical_compound ,Pituitary Gland, Anterior ,Endopeptidases ,Animals ,Pyroglutamyl-peptidase I ,Deamidation ,Thyrotropin-Releasing Hormone ,chemistry.chemical_classification ,Dipeptide ,Catabolism ,Rats, Inbred Strains ,Peptide Fragments ,Pyrrolidonecarboxylic Acid ,Rats ,Enzyme ,chemistry ,Chromatography, Gel - Abstract
Rapid fragmentation of thyroliberin (less than Glu-His-Pro-NH2) by rat adenohypophyseal tissue enzymes could be demonstrated. Based on the identification of the metabolic products and by the demonstration that the individual enzymatic reactions can be preferentially blocked by enzyme inhibitors, specific and sensitive biochemical tests could be developed in order to monitor the enzymatic activities after gel chromatographic fractionation of the tissue extracts. These findings are in agreement with the interpretation that the observed degradation of thyroliberin by hypophyseal tissue extracts may follow the proposed pathways. The primary enzymatic cleavage of thyroliberin is either initiated by the action of a 'thyroliberin-deamidating enzyme' (thyroliberin leads to less than Glu-His-Pro-OH + NH3), or by the action of a pyroglutamate aminopeptidase (thyroliberin leads to less than Glu + His-Pro-NH2). While the pyroglutamate aminopeptidase also catalyzes the subsequent degradation of deamidated thyroliberin (less than Glu-His-Pro-OH leads to less than Glu + His-Pro-OH), the enzymatic deamidation of His-Pro-NH2 is not catalyzed by the 'thyroliberin-deamidating enzyme; but by a post-proline dipeptidyl aminopeptidase. Hydrolysis of the common intermediary metabolite His-Pro-OH to the free amino acids is apparently catalyzed by a proline dipeptidase. In addition to these enzymatic events rapid cyclization of His-Pro-NH2 to histidyl-proline-diketopiperazine His-Pro could be observed. This reaction however is mainly due to the non-enzymatic intramolecular condensation reaction which is characteristic for proline-containing dipeptide derivatives. An enzymatic activity which catalyzes this reaction could not be observed when the enzyme fractions were tested. Enzymatic degradation of His-Pro by hypophyseal tissue extracts could also not be observed.
- Published
- 2005
- Full Text
- View/download PDF
4. Enzymatische Synthese biologisch aktiver Antibiotikapeptide und immunologisch suppressiver Cyclosporinderivate. Reaktive Sauerstoffspezies: Prooxidantien und Antioxidantien in Biologie und Medizin : 380. Sitzung am 9. Januar 1992 in Düsseldorf
- Author
-
Horst Kleinkauf and Horst Kleinkauf
- Subjects
- Cyclosporine, Peptides--Synthesis, Cyclosporins--biosynthesis, Oxidants, Peptides--biosynthesis
- Published
- 2013
5. Directed Biosynthesis of New Enniatins
- Author
-
Antje Lindemann, Wolfgang Gau, Mirko Gnnski, Till Hornbogen, Horst Kleinkauf, Gerhard Tmelking, Rainer Zocher, Peter Jeschke, Gerhard Bonse, and Martin Krause
- Subjects
Magnetic Resonance Spectroscopy ,Stereochemistry ,Mass Spectrometry ,Fungal Proteins ,Serine ,chemistry.chemical_compound ,Fusarium ,Biosynthesis ,Depsipeptides ,Drug Discovery ,Peptide Synthases ,Threonine ,Chromatography, High Pressure Liquid ,Anthelmintics ,Pharmacology ,chemistry.chemical_classification ,Alanine ,Depsipeptide ,Anti-Bacterial Agents ,Culture Media ,Amino acid ,chemistry ,Biochemistry ,Chromatography, Thin Layer ,Peptides ,Enniatin ,Cysteine - Abstract
New cyclohexadepsipeptides of the enniatin type with potential anthelmintic properties were produced by two different strategies: 1. In vitro synthesis by use of the multienzyme enniatin synthetase, and 2. in vivo precursor feeding of enniatin producing strains Fusarium scirpi and Fusarium sambucinum. The compounds were analyzed by HPLC, various NMR measurements and mass spectrometry. The three N-methyl L-amino acid positions in the enniatin B molecule could be gradually replaced by other (N-methyl) L-amino acids, e.g. alanine, cysteine, threonine and serine. The latter two amino acids yield new enniatins with functional groups in the hydrophobic side chains. Similarly the three D-2-hydroxyisovalerate residues, present in all naturally occuring enniatins, could be substituted by D-2-hydroxybutyric acid and D-lactic acid. Despite its lower yield the in vitro synthesis has the advantage of a broader variety of products formed.
- Published
- 2001
- Full Text
- View/download PDF
6. Biosynthesis of PF1022A and Related Cyclooctadepsipeptides
- Author
-
Wolfram Weckwerth, Koichi Miyamoto, Katsuhura Iinuma, Martin Krause, Mirko Glinski, Thomas Storm, Gerd Bonse, Horst Kleinkauf, and Rainer Zocher
- Subjects
Protein Conformation ,Stereochemistry ,Peptide ,Biology ,Peptides, Cyclic ,Biochemistry ,chemistry.chemical_compound ,Adenosine Triphosphate ,Protein structure ,Biosynthesis ,Depsipeptides ,Peptide bond ,Amino Acids ,Peptide Synthases ,Binding site ,Molecular Biology ,Adenylylation ,Chromatography, High Pressure Liquid ,chemistry.chemical_classification ,Depsipeptide ,Chromatography ,Fungi ,Cell Biology ,Kinetics ,Enzyme ,chemistry - Abstract
PF1022A belongs to a recently identified class of N-methylated cyclooctadepsipeptides (CODPs) with strong anthelmintic properties. Described here is the cell-free synthesis of this CODP and related structures, as well as the purification and enzymatic characterization of the responsible synthetase. For PF1022A synthesis extracts of Mycelia sterilia were incubated with the precursors L-leucine, D-lactate, D-phenyllactate, and S-adenosyl-L-methionine in the presence of ATP and MgCl(2). A 350-kDa depsipeptide synthetase, PFSYN, responsible for PF1022A synthesis was purified to electrophoretic homogeneity. Like other peptide synthetases, PFSYN follows a thiotemplate mechanism in which the substrates are activated as thioesters via adenylation. N-Methylation of the substrate L-leucine takes place after covalent binding prior to peptide bond formation. The enzyme is capable of synthesizing all known natural cyclooctadepsipeptides of the PF1022 type (A, B, C, and D) differing in the content of D-lactate and D-phenyllactate. In addition to PF1022 types A, B, C, and D, the in vitro incubations produced PF1022F (a CODP consisting of D-lactate and N-methyl-L-leucine), as well as di-, tetra-, and hexa-PF1022 homologs. PFSYN strongly resembles the well documented enniatin synthetase in size and mechanism. Our results suggest that PFSYN, like enniatin synthetase, is an enzyme with two peptide synthetase domains and forms CODP by repeated condensation of dipeptidol building blocks. Due to the low specificity of the d-hydroxy acid binding site, D-lactate or D-phenyllactate can be incorporated into the dipeptidols depending on the concentration of these substrates in the reaction mixture.
- Published
- 2000
- Full Text
- View/download PDF
7. Editing of non-cognate aminoacyl adenylates by peptide synthetases
- Author
-
Horst Kleinkauf, Ralf Dieckmann, Maja Pavela-Vrančić, and Hans von Döhren
- Subjects
Stereochemistry ,Adenylate kinase ,Bacillus ,Peptide ,Biology ,Biochemistry ,Pyrophosphate ,Catalysis ,Hydrolysis ,chemistry.chemical_compound ,Adenosine Triphosphate ,Escherichia coli ,heterocyclic compounds ,Nucleotide ,Peptide Synthases ,Pyrophosphatases ,Molecular Biology ,Adenylylation ,chemistry.chemical_classification ,non-ribosomal ,proof reading ,Substrate (chemistry) ,Cell Biology ,Adenosine Monophosphate ,Amino acid ,chemistry ,Apoproteins ,Research Article - Abstract
Non-ribosomally formed peptides display both highly conserved and variable amino acid positions, the variations leading to a wide range of peptide families. Activation of the amino acid substrate proceeds in analogy to the ribosomal biosynthetic mechanism generating aminoacyl adenylate and acyl intermediates. To approach the mechanism of fidelity of amino acid selection, the stability of the aminoacyl adenylates was studied by employing a continuous coupled spectrophotometric assay. The apo-form of tyrocidine synthetase 1 (apo-TY1) was used, generating an L-phenylalanyl-adenylate intermediate stabilized by the interaction of two structural subdomains of the adenylation domain. Adenylates of substrate analogues have shown variable and reduced degrees of stability, thus leading to an enhanced generation of pyrophosphate due to hydrolysis and continuous adenylate formation. Discrimination of the non-aromatic amino acids L-Leu and L-Met, or L-Phe analogues such as p-amino- and p-chloro-L-Phe derivatives, as well as the stereospecific selection of L-Phe, is supported by less-stable adenylate intermediates exhibiting elevated susceptibility to hydrolysis. Breakdown of the L-phenylalanyl intermediate utilizing 2′-deoxy-ATP as the nucleotide substrate was significantly enhanced compared with the natural analogue. Apo-TY1 engineered at positions involved in adenylate formation showed variable protection against hydrolysis. The results imply that stability of the aminoacyl intermediates may act as an essential factor in substrate selection and fidelity of non-ribosomal-peptide-forming systems.
- Published
- 1999
- Full Text
- View/download PDF
8. Probing the domain structure and ligand-induced conformational changes by limited proteolysis of tyrocidine synthetase 1
- Author
-
Horst Kleinkauf, Maja Pavela-Vrančić, Ralf Dieckmann, and Hans von Döhren
- Subjects
Protein Folding ,Protein Conformation ,Sequence analysis ,Stereochemistry ,Proteolysis ,Adenylyl Imidodiphosphate ,Peptide ,Multifunctional Enzymes ,Ligands ,Cleavage (embryo) ,Catalysis ,Adenosine Triphosphate ,Allosteric Regulation ,Structural Biology ,medicine ,Luciferase ,Peptide Synthases ,Molecular Biology ,Adenylylation ,Sequence Deletion ,chemistry.chemical_classification ,Binding Sites ,Peptide synthetases ,limited proteolysis ,domain organization ,linker ,medicine.diagnostic_test ,Chemistry ,Peptide Fragments ,Diphosphates ,Kinetics ,Linker ,Allosteric Site - Abstract
The boundaries of the structural domains in peptide synthetases and the conformational changes related to catalysis were investigated by limited proteolysis of tyrocidine synthetase 1 (TY1). Four regions sensitive to proteolysis were detected (cleavage site at Arg13, Arg424, Arg509 and Arg602) that, in addition to an N-terminal extension, accurately delineate the domain boundaries of the adenylate-forming domain, the aminoacyl carrier domain, and the epimerisation domain. Limited proteolysis of an active N-terminal truncated deletion mutant, His6ΔTY1, generated two stable and structurally independent subunits, corresponding to the subdomains of the adenylation domain. The structural integrity of the carrier domain was substantiated by its resistance to proteolytic degradation. Evidence is provided that the C-terminal “spacer” region with epimerising and/or condensing activity folds into an autonomous domain stable against degradation by limited proteoly sis. In the presence of substrates, reduced susceptibility to proteolysis was observed in the linker region connecting the subdomains of the adenylation domain, and corresponding to a peptide stretch of low electron density in the X-ray structure of the homologous firefly luciferase. Sequence analysis has shown that the respective linker contains conserved residues, whereas the linker regions connecting the structural domains are of low homology with a significant content of Pro, Ala, Glu and polar residues. A combination of kinetic and proteolytic studies using ATP analogues with substitutions in the phosphate chain, AMP-PcP, AMP-P n P and AMP-cPP, strongly suggests that the generation of a productive complex is associated with the ability of the β,γ-pyrophosphate moiety of ATP to adopt the proper active-site conformation. These data substantiate the observation that peptide synthetases undergo a series of conformational changes in the process of adenylate formation and product release.
- Published
- 1999
- Full Text
- View/download PDF
9. Progress in Drug Research / Fortschritte Der Arzneimittelforschung / Progrès Des Recherches Pharmaceutiques
- Author
-
Eric J. Lien, Arima Das, Partha Nandy, Shijun Ren, Horst Kleinkauf, Hans von Döhren, Iradj Hajimohamadreza, J. Mark Treherne, Esteban Domingo, Luis Menéndez-Arias, Miguel E. Quiñones-Mateu, Africa Holguín, Mónica Gutiérrez-Rivas, Miguel A. Martínez, Josep Quer, Isabel S. Novella, John J. Holland, Vijendra K. Singh, Deborah S. Hartman, Olivier Civelli, Vera M. Kolb, Eric J. Lien, Arima Das, Partha Nandy, Shijun Ren, Horst Kleinkauf, Hans von Döhren, Iradj Hajimohamadreza, J. Mark Treherne, Esteban Domingo, Luis Menéndez-Arias, Miguel E. Quiñones-Mateu, Africa Holguín, Mónica Gutiérrez-Rivas, Miguel A. Martínez, Josep Quer, Isabel S. Novella, John J. Holland, Vijendra K. Singh, Deborah S. Hartman, Olivier Civelli, and Vera M. Kolb
- Subjects
- Pharmacology
- Published
- 2012
10. Applications of peptide synthetases in the synthesis of peptide analogues
- Author
-
H von Döhren and Horst Kleinkauf
- Subjects
chemistry.chemical_classification ,Amino acid activation ,Peptide ,General Biochemistry, Genetics and Molecular Biology ,chemistry.chemical_compound ,chemistry ,Biochemistry ,Tyrocidine ,polycyclic compounds ,Gramicidin ,Peptide synthesis ,Peptide Biosynthesis ,Peptide library ,Peptide sequence - Abstract
Enzymatically formed peptides show positional variations as well as highly conserved amino acids. In the cases of gramicidin S, tyrocidine, linear gramicidins, enniatins, echinocandins and viridogrisein in vivo and in vitro studies indicate substrate selection at the level of amino acid activation as a major control step. Evidence for proof-reading steps beyond activation has been obtained in penicillin and cyclosporin biosynthesis. Activated substrate analogues may promote the formation of side products such as dipeptides and cyclodipeptides. Modifications of intermediates, such as N-methylation, influence the rates of peptide synthesis. These control steps pose limitations for the application of such enzyme systems in the production of peptide libraries. They may originate from a target oriented evolution of these synthetases.
- Published
- 1997
- Full Text
- View/download PDF
11. Reversible denaturation of cyclosporin synthetase by urea
- Author
-
Alfons Lawen, François Vaillant, Horst Kleinkauf, and J. Dittmann
- Subjects
Protein Denaturation ,Protein Folding ,Conformational change ,Protein Conformation ,Biophysics ,Biosynthesis ,Peptides, Cyclic ,Cyclosporin synthetase ,Biochemistry ,Piperazines ,chemistry.chemical_compound ,Protein structure ,Multienzyme Complexes ,Structural Biology ,Cyclosporin a ,Genetics ,Urea ,Denaturation (biochemistry) ,Peptide Synthases ,Molecular Biology ,chemistry.chemical_classification ,cyclo-(d-alanyl-N-methylleucyl) ,Fungi ,Folding ,Cell Biology ,Enzyme ,Fluorescence spectrum ,chemistry ,Cyclosporine ,Protein folding ,Multidomain enzyme - Abstract
The reversible denaturation of the multifunctional polypeptide, cyclosporin synthetase, by urea was analyzed. It is possible to discriminate between at least two stages of enzyme denaturation. While at low urea concentration (up to 0.8 M) cyclosporin A formation is inhibited, synthesis of the diketopiperazine cyclo-(d-alanyl-N-methylleucyl), a molecule representing a partial sequence of cyclosporin A is still detectable. At higher concentrations of urea the enzyme preparation is totally inactive. This inactivation is a consequence of conformational change(s) of cyclosporin synthetase as shown by fluorescence emission spectra of native and denatured enzyme. These data imply a consecutive folding/defolding mechanism for the different domains forming the multifunctional polypeptide.
- Published
- 1996
- Full Text
- View/download PDF
12. Characterization of tyrocidine synthetase 1 (TY1): Requirement of posttranslational modification for peptide biosynthesis
- Author
-
Maja Pavela-Vrančić, Horst Kleinkauf, Eva Pfeifer, and Von Doehren H
- Subjects
Peptide Biosynthesis ,Molecular Sequence Data ,Bacillus ,Aminoacylation ,Biochemistry ,Mass Spectrometry ,Substrate Specificity ,chemistry.chemical_compound ,Adenosine Triphosphate ,Escherichia coli ,Serine ,Peptide synthesis ,Amino Acid Sequence ,Cloning, Molecular ,Peptide Synthases ,Adenylylation ,Conserved Sequence ,chemistry.chemical_classification ,Binding Sites ,Sequence Homology, Amino Acid ,Molecular mass ,Chemistry ,Phosphopantetheine binding ,Peptide Fragments ,Recombinant Proteins ,Amino acid ,Molecular Weight ,Kinetics ,Genes, Bacterial ,Phosphopantetheine ,Peptides ,Protein Processing, Post-Translational - Abstract
Tyrocidine synthetase 1 (TY1), produced by Bacillus brevis ATCC 8185, consists of a single multifunctional polypeptide chain catalyzing the activation, thioesterification, and epimerization of phenylalanine. Because we were concerned about possible posttranslational issues, a comparative study between the wild-type isolate and the in Escherichia coli overexpressed protein was performed. Analysis by matrix assisted laser desorption mass spectrometry (MALDI) provided a molecular mass of 122,516 +/- 120 Da for the recombinant protein, which is in agreement with the value of 122,590 Da calculated from the gene sequence. MALDI analysis of the tryptic fragments revealed that in the recombinant TY1 the putative 4'-phosphopantetheine binding site (562Ser) is not modified by the cofactor. The substrate specificity profiles of the amino acid dependent ATP[32P]PPi exchange reactions were identical, including activation of L-phenylserine, L-tyrosine, and L-methionine. However, the rates of the reverse adenylation reaction for the recombinant protein were only 22% relative to those of the wild-type enzyme. The aminoacylation levels of about 60% for TY1 from Bacillus brevis reduced to 1.4% in the overexpressed protein. A similar distribution of the D- and the L-isomer was detected at the thioester attachment site. The pI values of the wild-type and expressed TY1 are 4.9 and 5.0, respectively. In conclusion, it could be established that apo- and holo-TY1 differ in their amino acid activating properties. Posttranslational modification by 4'-phosphopantetheine is an essential requirement for aminoacylation, epimerization, and thus the functioning of the multienzyme in peptide synthesis.
- Published
- 1995
- Full Text
- View/download PDF
13. ATP Binding in Peptide Synthetases: Determination of Contact Sites of the Adenine Moiety by Photoaffinity Labeling of Tyrocidine Synthetase 1 with 2-Azidoadenosine Triphosphate
- Author
-
Hans-Jochen Schaefer, Maja Pavela-Vrančić, Horst Kleinkauf, Eva Pfeifer, Hans von Doehren, and Henk van Liempt
- Subjects
Azides ,Photochemistry ,Ultraviolet Rays ,Sequence analysis ,Stereochemistry ,Biochemistry ,Adenosine Triphosphate ,Ultraviolet light ,Trypsin ,Nucleotide ,Peptide Synthases ,Chromatography, High Pressure Liquid ,Amino acid activation ,chemistry.chemical_classification ,Binding Sites ,Photoaffinity labeling ,Chemistry ,Adenine ,Protein primary structure ,Affinity Labels ,Peptide Fragments ,Kinetics ,Enzyme ,Cyclic nucleotide-binding domain ,Chromatography, Gel - Abstract
Characterization of the nucleotide binding domain in peptide synthetases was approached by photoaffinity labeling of tyrocidine synthetase 1 (TY1) with 2-azidoadenosine triphosphate (2-azido-ATP). Exposure of TY1 in the presence of photolabel to irradiation with ultraviolet light resulted in a time-dependent covalent modification of the enzyme with a concomitant loss of catalytic activity. Inactivation was not observed if incubation was performed in the absence of either light or the nucleotide analogue. Specificity of labeling was indicated by the ability of 2-azido-ATP to serve as a substrate in the amino acid activation reaction. The modified protein was subjected to tryptic digestion, and the fragments labeled by the nucleotide analogue were purified by reverse-phase high-performance liquid chromatography. Sequence analysis identified three tryptic peptides corresponding to residues G373-K384, W405-R416, and L483-K494, derived from the N-terminal half of the TY1 sequence. As this region shows similarity to strongly conserved regions in other peptide synthetases and acyl-CoA synthetases, it is considered to be the region catalyzing aminoacyl adenylate formation. The identified sequences appear to define components of the nucleotide binding domain found in close proximity to the adenine ring in ATP. Conservation of primary structure and homology to other carboxyl-activating enzymes of this superfamily, including peptide synthetases, insect luciferases, and acyl-CoA synthetases, is discussed.
- Published
- 1994
- Full Text
- View/download PDF
14. Purification and characterization of eucaryotic alanine racemase acting as key enzyme in cyclosporin biosynthesis
- Author
-
Rainer Zocher, Horst Kleinkauf, E Schneider-Scherzer, and Kai Hoffmann
- Subjects
Stereochemistry ,Biology ,Biochemistry ,Piperazines ,chemistry.chemical_compound ,Multienzyme Complexes ,Cyclosporin a ,Alanine racemase ,Arginine racemase ,Peptide Synthases ,Pyridoxal phosphate ,Molecular Biology ,Alanine ,Gel electrophoresis ,chemistry.chemical_classification ,Molecular mass ,Alanine Racemase ,Cell Biology ,Enzyme ,chemistry ,Chromatography, Gel ,Cyclosporine ,Electrophoresis, Polyacrylamide Gel ,Chromatography, Thin Layer ,Mitosporic Fungi - Abstract
A specific alanine racemase, which is a key enzyme in the biosynthesis of the undecapeptide cyclosporin A, was purified to electrophoretic homogeneity from the fungus Tolypocladium niveum. This is the first enzyme of this kind isolated from a eucaryotic organism. The enzyme catalyzes the reversible racemization of alanine and requires pyridoxal phosphate as the exclusive cofactor. Km values for L- and D-alanine were found to be 38 and 2 mM, respectively. Maximal reaction velocity was observed at 42 degrees C and pH 8.8 for the L to D direction. Molecular mass determinations of the denatured enzyme by SDS-polyacrylamide gel electrophoresis gave a value of 37 kDa, whereas gel filtration calibration studies yielded a value between 120 and 150 kDa, indicating an oligomeric native structure.
- Published
- 1994
- Full Text
- View/download PDF
15. Mechanism of cyclosporin A biosynthesis. Evidence for synthesis via a single linear undecapeptide precursor
- Author
-
R.M. Wenger, Horst Kleinkauf, J. Dittmann, and Alfons Lawen
- Subjects
chemistry.chemical_classification ,Stereochemistry ,Peptide ,Cell Biology ,Thioester ,Biochemistry ,Amino acid ,chemistry.chemical_compound ,Biosynthesis ,chemistry ,Cyclosporin a ,Biosynthetic process ,polycyclic compounds ,Peptide bond ,Molecular Biology ,Peptide sequence - Abstract
Cyclosporin A is synthesized by cyclosporin synthetase, a multienzyme polypeptide. This enzyme catalyzes at least 40 reaction steps in an assembly belt-like mechanism. It activates all constituent amino acids of cyclosporin A to thioesters via amino acyladenylates and carries out specific N-methylation reactions. During elongation, the activated amino acids are linked by peptide bonds leading to enzyme-bound nascent peptide chains. Some of the linear peptides of the growing cyclosporin A chain were isolated and their N-terminal amino acid was determined. D-Alanine at position 8 of the cyclosporin A molecule was found to be a starting amino acid in the biosynthetic process of cyclosporin A formation. Four intermediate peptides of the growing peptide chain of cyclosporin A could be isolated and identified. All of them represent partial sequences of cyclosporin A starting with D-alanine. That these intermediate peptides were bound by thioester linkage to cyclosporin synthetase could be demonstrated by liberation of the peptides with performic acid. The peptides strongly suggest the stepwise synthesis of a single linear peptide precursor of cyclosporin A.
- Published
- 1994
- Full Text
- View/download PDF
16. 4.1 The Squiggle - Symbol of Bioenergetics
- Author
-
Horst Kleinkauf, Lothar Jaenicke, and Hans von Döhren
- Subjects
Symbol ,Communication ,business.industry ,media_common.quotation_subject ,business ,media_common ,Mathematics - Published
- 2011
- Full Text
- View/download PDF
17. 1. Fritz Lipmann (1899-1986)
- Author
-
Horst Kleinkauf, Lothar Jaenicke, and Hans von Döhren
- Published
- 2011
- Full Text
- View/download PDF
18. 2. Biochemistry Comes of Age
- Author
-
Horst Kleinkauf, Lothar Jaenicke, and Hans von Döhren
- Subjects
Physiology ,Biology - Published
- 2011
- Full Text
- View/download PDF
19. Fritz Lipmann: Bibliography 1924-1985
- Author
-
Horst Kleinkauf, Lothar Jaenicke, and Hans von Döhren
- Subjects
Philosophy ,Bibliography ,Classics - Published
- 2011
- Full Text
- View/download PDF
20. 4.2 Molecular Recognition and Communication
- Author
-
Hans von Döhren, Lothar Jaenicke, and Horst Kleinkauf
- Subjects
Molecular recognition ,Computer science ,Computational biology - Published
- 2011
- Full Text
- View/download PDF
21. 3. Molecular Biology Sharpens its Tools
- Author
-
Lothar Jaenicke, Horst Kleinkauf, and Hans von Döhren
- Subjects
Computational biology ,Biology - Published
- 2011
- Full Text
- View/download PDF
22. Research on Nonribosomal Systems: Biosynthesis of Peptide Antibiotics
- Author
-
Hans von Döhren and Horst Kleinkauf
- Published
- 2011
- Full Text
- View/download PDF
23. ChemInform Abstract: Biosynthesis of Peptides: A Nonribosomal System
- Author
-
H. Von Doehren, Horst Kleinkauf, H. Van Liempt, and H. Palissa
- Subjects
chemistry.chemical_compound ,Biochemistry ,Biosynthesis ,Chemistry ,General Medicine - Published
- 2010
- Full Text
- View/download PDF
24. ChemInform Abstract: Linking Peptide and Polyketide Biosynthesis
- Author
-
Horst Kleinkauf and H. Von Doehren
- Subjects
chemistry.chemical_classification ,Biochemistry ,chemistry ,Polyketide biosynthesis ,Peptide ,General Medicine - Published
- 2010
- Full Text
- View/download PDF
25. ChemInform Abstract: Applications of Peptide Synthetases in the Synthesis of Peptide Analogues
- Author
-
H von Döhren and Horst Kleinkauf
- Subjects
Amino acid activation ,chemistry.chemical_classification ,Peptide ,General Medicine ,Amino acid ,chemistry.chemical_compound ,chemistry ,Biosynthesis ,Biochemistry ,Tyrocidine ,polycyclic compounds ,Peptide synthesis ,Gramicidin ,Echinocandins - Abstract
Enzymatically formed peptides show positional variations as well as highly conserved amino acids. In the cases of gramicidin S, tyrocidine, linear gramicidins, enniatins, echinocandins and viridogrisein in vivo and in vitro studies indicate substrate selection at the level of amino acid activation as a major control step. Evidence for proof-reading steps beyond activation has been obtained in penicillin and cyclosporin biosynthesis. Activated substrate analogues may promote the formation of side products such as dipeptides and cyclodipeptides. Modifications of intermediates, such as N-methylation, influence the rates of peptide synthesis. These control steps pose limitations for the application of such enzyme systems in the production of peptide libraries. They may originate from a target oriented evolution of these synthetases.
- Published
- 2010
- Full Text
- View/download PDF
26. ChemInform Abstract: Directed Biosynthesis of New Enniatins
- Author
-
Mirko Glinski, Peter Jeschke, Wolfgang Gau, Rainer Zocher, Horst Kleinkauf, Martin Krause, Gerhard Thielking, Gerhard Bonse, Till Hornbogen, and Antje Lindemann
- Subjects
chemistry.chemical_classification ,chemistry.chemical_compound ,chemistry ,Biosynthesis ,Biochemistry ,General Medicine ,Amino acid - Published
- 2010
- Full Text
- View/download PDF
27. Two new cyclophilins from Fusarium sambucinum and Aspergillus niger: Resistance of cyclophilin/cyclosporin a complexes against proteolysis
- Author
-
Till Hornbogen, Kai Hoffmann, Horst Kleinkauf, Rainer Zocher, and Rembert Pieper
- Subjects
Isomerase activity ,Molecular Sequence Data ,Cyclosporin A binding ,Biophysics ,Biochemistry ,Microbiology ,Fusarium ,Tolypocladium inflatum ,Cyclosporin a ,Endopeptidases ,polycyclic compounds ,Chymotrypsin ,Amino Acid Sequence ,Molecular Biology ,Chromatography, High Pressure Liquid ,Cyclophilin ,Amino Acid Isomerases ,Peptidylprolyl isomerase ,Chromatography ,biology ,Serine Endopeptidases ,Aspergillus niger ,Proteolytic enzymes ,Metalloendopeptidases ,Cell Biology ,Peptidylprolyl Isomerase ,biology.organism_classification ,Molecular Weight ,enzymes and coenzymes (carbohydrates) ,Cyclosporine ,cardiovascular system ,Carrier Proteins - Abstract
Two new peptidyl-prolyl-cis/trans-isomerases were purified to homogeneity from Fusarium sambucinum and Aspergillus niger . They belong to the class of cyclosporin A binding proteins (cyclophilins) and have molecular masses of about 18 kDa. As has been shown for other cyclophilins, the isomerase activity of the enzymes is inhibited by cyclosporin A in the nanomolar range. Furthermore binding of cyclosporin A prevents proteolytic digestion of the cyclophilin/cyclosporin complexes by the endoproteases GluC, LysC and α-chymotrypsin, in contrast to the free cyclophilins, which are readily cleaved by these proteases. We could also observe this protection for cyclophilins from sheep thymus and from the cyclosporin producing fungus Tolypocladium inflatum .
- Published
- 1992
- Full Text
- View/download PDF
28. A highly specific D-hydroxyisovalerate dehydrogenase from the enniatin producer Fusarium sambucinum
- Author
-
Rainer Zocher, Horst Kleinkauf, Helmut Görisch, and Chan Lee
- Subjects
Stereochemistry ,Dehydrogenase ,Peptides, Cyclic ,Biochemistry ,Redox ,Michaelis–Menten kinetics ,Catalysis ,Reversible reaction ,Substrate Specificity ,Hemiterpenes ,Fusarium ,Depsipeptides ,Peptide Synthases ,Molecular Biology ,chemistry.chemical_classification ,biology ,Molecular mass ,Temperature ,Cell Biology ,Fungi imperfecti ,Hydrogen-Ion Concentration ,NAD ,biology.organism_classification ,Keto Acids ,Anti-Bacterial Agents ,Molecular Weight ,Alcohol Oxidoreductases ,Kinetics ,Enzyme ,chemistry ,Chromatography, Gel ,Autoradiography ,Electrophoresis, Polyacrylamide Gel ,Peptides ,Enniatin ,Oxidation-Reduction ,NADP - Abstract
A highly specific D-hydroxyisovalerate (D-HIV) dehydrogenase, which is a key enzyme in depsipeptide synthesis, was purified to near homogeneity from the enniatin-producing fungus Fusarium sambucinum. The enzyme catalyzes the reversible reaction of 2-ketoisovalerate (2-KIV) to D-HIV. It is strictly dependent on NADPH and exhibits a high substrate specificity with respect to 2-KIV. NADH was not accepted by the enzyme. Km values for 2-KIV and NADPH were found to be 200 and 333 microM, respectively. D-HIV dehydrogenase consists of a single polypeptide chain with a molecular mass of about 53 kDa. Optimum temperature for the reduction of 2-KIV was 35 degrees C and for the oxidation reaction was 45 degrees C. The optimum pH was found to be 7 for the reduction and 8-9 for the oxidation reaction.
- Published
- 1992
- Full Text
- View/download PDF
29. Biosynthese von Peptiden: Ein nichtribosomales System
- Author
-
H van Liempt, Horst Kleinkauf, H. Palissa, and H von Döhren
- Subjects
chemistry.chemical_classification ,Natural product ,Chemistry ,Stereochemistry ,Microbial metabolism ,Peptide ,General Medicine ,chemistry.chemical_compound ,Biochemistry ,Biosynthesis ,Nucleic acid ,Peptide Biosynthesis ,Peptide sequence ,Ecology, Evolution, Behavior and Systematics ,Macromolecule - Abstract
The biosynthesis of peptides in nonribosomal systems is accomplished by complex multienzymes. These multienzymes assemble the required template for the construction of each natural product in the form of linearly coupled modules. This organization principle permits the integration of multistep synthetic processes on a single macromolecule.
- Published
- 1992
- Full Text
- View/download PDF
30. Enzymatic characterisation of the multifunctional enzyme delta-(l-alpha-aminoadipyl)-l-cysteinyl-d-valine synthetase from Streptomyces clavuligerus
- Author
-
Henk van Liempt, Horst Kleinkauf, H. Palissa, Torsten Schwecke, Hans von Döhren, and Yair Aharonowitz
- Subjects
chemistry.chemical_classification ,Gel electrophoresis ,biology ,Molecular mass ,Dithioerythritol ,Stereochemistry ,ACV synthetase ,Streptomyces clavuligerus ,Chromatography, Ion Exchange ,biology.organism_classification ,Biochemistry ,Streptomyces ,Amino acid ,Amino Acyl-tRNA Synthetases ,Molecular Weight ,Kinetics ,chemistry.chemical_compound ,Enzyme ,chemistry ,Valine ,Chromatography, Gel ,biology.protein ,Electrophoresis, Polyacrylamide Gel ,Peptide Synthases - Abstract
delta-(L-alpha-Aminoadipyl)-L-cysteinyl-D-valine (ACV) synthetase, the multienzyme catalyzing the formation of ACV from the constituent amino acids and ATP in the presence of Mg2+ and dithioerythritol, was purified about 2700-fold from Streptomyces clavuligerus. The molecular mass of the native enzyme as determined by gel filtration chromatography is 560 kDa, while that determined by denaturing gel electrophoresis is 500 kDa. The enzyme is able to catalyze pyrophosphate exchange in dependence on L-cysteine and L-valine, but no L-alpha-aminoadipic-acid-dependent ATP/PPi exchange could be detected. Other L-cysteine- and L-valine-activating enzymes present in crude extracts were identified as aminoacyl-tRNA synthetases which could be separated from ACV synthetase. The molecular mass of these enzymes is 140 kDa for L-valine ligase and 50 kDa for L-cysteine ligase. The dissociation constants have been estimated, assuming three independent activation sites, to be 1.25 mM and 1.5 mM for cysteine and ATP, and 2.4 mM and 0.25 mM for valine and ATP, respectively. The enzyme forms a thioester with alpha-aminoadipic acid and with valine in a molar ratio of 0.6:1 (amino acid/enzyme). Thus, the bacterial ACV synthetase is a multifunctional peptide synthetase, differing from fungal ACV synthetases in its mechanism of activation of the non-protein amino acid.
- Published
- 1992
- Full Text
- View/download PDF
31. Enniatin synthetases from different fusaria exhibiting distinct amino acid specificities
- Author
-
Horst Kleinkauf, Rainer Zocher, and Rembert Pieper
- Subjects
Pharmacology ,chemistry.chemical_classification ,DNA ligase ,biology ,Stereochemistry ,Fusarium sambucinum ,Antibodies, Monoclonal ,Fungi imperfecti ,biology.organism_classification ,Substrate Specificity ,Amino acid ,Enzyme ,Fusarium ,Biochemistry ,chemistry ,Fusarium lateritium ,Drug Discovery ,Fusarium scirpi ,Peptide Synthases ,Enniatin - Abstract
Enniatin synthetases from the cyclodepsipeptide producers Fusarium lateritium and Fusarium sambucinum were purified to homogeneity and characterized. Like the previously described enniatin synthetase from Fusarium scirpi both enzymes consist of a single polypeptide chain and are very similar concerning their Mr (250 kdaltons) and reaction mechanism. Limited proteolytic digests show only slight differences in their patterns in SDS-gels. Interestingly the synthetases differ in their amino acids specificities. The enzyme from the enniatin A producer F. sambucinum exhibits a high affinity to the substrate amino acids L-Leu and L-Ile. In contrast the synthetase from the enniatin B producer F. lateritium preferably accepts L-Val, the constituent amino acid of enniatin B.
- Published
- 1992
- Full Text
- View/download PDF
32. Localization of isopenicillin N synthase in Penicillium chrysogenum PQ-96
- Author
-
H. Von Dohren, Horst Kleinkauf, W. Kuryłowicz, W. P. Wolf, H. Van Liempt, W. Kurzătkowski, and H. Palissa
- Subjects
biology ,Isopenicillin N synthase ,General Medicine ,Golgi apparatus ,Penicillium chrysogenum ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Cell wall ,symbols.namesake ,Biochemistry ,Labelling ,Lowicryl K4M ,Cephalosporium acremonium ,biology.protein ,symbols ,Mycelium ,Biotechnology - Abstract
For immunoelectron microscopic localization of isopenicillin N synthase (IPNS), glutaraldehyde-fixed mycelium of Penicillium chrysogenum PQ-96 was dehydrated by progressive lowering of the temperature and embedded into Lowicryl K4M at −35° C. This procedure resulted in good structural preservation such that the method of on-section labelling using antibody to IPNS from Cephalosporium acremonium CO 278 with the indirect antibody-gold technique could be successfully applied. By this method IPNS was localized in vesicular compartments belonging probably to the Golgi body and in the cell wall.
- Published
- 1991
- Full Text
- View/download PDF
33. Peptide Antibiotics
- Author
-
Horst Kleinkauf and Hans von Döhren
- Published
- 2008
- Full Text
- View/download PDF
34. Polypeptide Synthesis on Protein Templates: The Enzymatic Synthesis of Gramicidin S And Tyrocidine
- Author
-
Robert Roskoski, Horst Kleinkauf, Fritz Lipmann, and Wieland Gevers
- Subjects
chemistry.chemical_compound ,Template ,Biochemistry ,Stereochemistry ,Chemistry ,Tyrocidine ,Tyrocidine biosynthesis ,Gramicidin S ,Enzymatic synthesis - Published
- 2006
- Full Text
- View/download PDF
35. New beauveriolides produced by amino acid-supplemented fermentation of Beauveria sp. FO-6979
- Author
-
Horst Kleinkauf, Susumu Kobayashi, Satoshi Omura, Daisuke Matsuda, Rainer Zocher, Ichiji Namatame, and Hiroshi Tomoda
- Subjects
Spectrophotometry, Infrared ,Stereochemistry ,Microbial Sensitivity Tests ,Spectrometry, Mass, Fast Atom Bombardment ,Peptides, Cyclic ,chemistry.chemical_compound ,Mice ,Ascomycota ,Lipid droplet ,Depsipeptides ,Drug Discovery ,Acetone ,Animals ,Amino Acids ,Nuclear Magnetic Resonance, Biomolecular ,Chemical decomposition ,Mycelium ,Pharmacology ,chemistry.chemical_classification ,Chromatography ,Molecular Structure ,Fatty acid ,General Medicine ,Lipid Metabolism ,Amino acid ,Anti-Bacterial Agents ,chemistry ,Fermentation ,Cholesteryl ester ,Chromatography, Gel ,Macrophages, Peritoneal ,Spectrophotometry, Ultraviolet ,Cholesterol Esters - Abstract
Five new beauveriolides were isolated from the acetone extracts of Beauveria sp. FO-6979 mycelia fermented in amino acid-supplemented media. The structures were elucidated by spectroscopic studies including NMR experiments and chemical degradation. All the beauveriolides are cyclodepsipeptides consisting of one 3-hydroxy-4-methyl fatty acid, two L-amino acids and one D-amino acid in common. Beauveriolide VII with the structure of cyclo-[3-hydroxy-4-methyloctanoyl-L-phenylalanyl-L-alanyl-D-valyl] inhibited lipid droplet formation and cholesteryl ester synthesis in macrophages, but the other beauveriolides showed only slight or almost no effect on lipid droplet formation.
- Published
- 2004
36. Peptides
- Author
-
Hans von Döhren and Horst Kleinkauf
- Published
- 2002
- Full Text
- View/download PDF
37. Thiotemplate Mechanism Of Peptide Antibiotic Synthesis
- Author
-
Horst Kleinkauf and Hans von Doehren
- Subjects
Peptide antibiotic synthesis ,Biochemistry ,Chemistry ,Mechanism (biology) ,human activities - Abstract
This article has no abstract. Keywords: thiotemplate mechanism of peptide antibiotic synthesis
- Published
- 2002
- Full Text
- View/download PDF
38. Peptide Antibiotic Biosynthesis
- Author
-
Horst Kleinkauf and Hans von Dozhren
- Subjects
Chemistry ,Peptide antibiotic biosynthesis ,Microbiology - Published
- 2002
- Full Text
- View/download PDF
39. ChemInform Abstract: Cyclodepsipeptides: From Natural Product to Anthelmintically Active Synthetic Enniatins
- Author
-
Peter Jeschke, Winfried Etzel, Achim Harder, Michael Schindler, Axel Gohrt, Ulrich Pleiss, Horst Kleinkauf, Rainer Zocher, Gerhard Thielking, Wolfgang Gau, and et al. et al.
- Subjects
chemistry.chemical_compound ,Natural product ,Chemistry ,Organic chemistry ,General Medicine ,Combinatorial chemistry - Published
- 2000
- Full Text
- View/download PDF
40. Cyclosporin: The Biosynthetic Path to a Lipopeptide
- Author
-
Horst Kleinkauf and Hans von Döhren
- Subjects
chemistry.chemical_compound ,chemistry ,Path (graph theory) ,Lipopeptide ,Topology - Published
- 1999
- Full Text
- View/download PDF
41. Penicillin biosynthesis: energy requirement for tripeptide precursor formation by delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Acremonium chrysogenum
- Author
-
H von Döhren, Horst Kleinkauf, and Wibke Dr. Kallow
- Subjects
Stereochemistry ,ACV synthetase ,Peptide ,Tripeptide ,Penicillins ,Biochemistry ,Peptide Synthases ,chemistry.chemical_compound ,Adenosine Triphosphate ,Nonribosomal peptide ,Enzyme Stability ,Peptide bond ,Amino Acids ,Chromatography, High Pressure Liquid ,chemistry.chemical_classification ,biology ,Chemistry ,Hydrolysis ,Amino acid ,Acremonium ,Dithiothreitol ,Kinetics ,Energy Transfer ,Reducing Agents ,biology.protein ,Electrophoresis, Polyacrylamide Gel ,Adenosine triphosphate ,Oligopeptides - Abstract
In nonribosomal peptide formation by multifunctional enzymes, peptide synthetases catalyze the activation and directed condensation of amino acids. The peptide synthetase involved in penicillin biosynthesis (ACV synthetase) forms the tripeptide delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine from the respective L-amino acids and ATP. So far, the energy requirements for the nonribosomal process have not been clearly established. For ACV synthetase we show that ATP consumption depends on the reaction conditions employed. By simultaneously estimating peptide and AMP production by employing fluorescence detection and UV spectroscopy, respectively, we have determined the energy consumption with high accuracy. Under unfavorable reaction conditions more than 20 mol of ATP are consumed/mol of tripeptide formed, while optimal conditions permit the expected energy requirement of one ATP for each carboxyl group activation, corresponding to three ATP for tripeptide formation. The third ATP is required for the activation of L-valine to maintain the valyl-thioester stage for epimerization and peptide bond formation, and this high-energy bond is sacrificed by hydrolytic removal of the product. No extra energy is required for the directed transport in peptide elongation. Additional energy consumed has been traced to hydrolytic loss of activated intermediates, as has been shown by the analysis of incomplete reaction mixtures.
- Published
- 1998
42. Enzymology of Peptide Synthetases
- Author
-
Hans von Döhren and Horst Kleinkauf
- Published
- 1997
- Full Text
- View/download PDF
43. Enzymatic generation of complex peptides
- Author
-
Horst Kleinkauf and H von Döhren
- Subjects
chemistry.chemical_classification ,chemistry.chemical_compound ,Protein structure ,chemistry ,Biochemistry ,Animal Sources ,Peptide synthesis ,Peptide ,Peptide Biosynthesis ,Pharmaceutical sciences ,Biology ,Bioproduction ,Cyclic peptide - Abstract
Complex peptides have continued to be lead structures in the search of drugs, especially those of microbial sources. In addition, the discovery of active peptides in animal sources including insects and mammals is advancing. As considerable progress has been achieved in the analysis of protein structure, the identification of targets and mechanisms of action is carried out at the molecular levels. For many purposes of structure-function studies the preparation and modification of complex peptides are required, and often chemical strategies are laborious or not available. Efforts have thus been conducted to make use of protein biocatalysts evolved in the producer organisms, exerting their catalytic powers under physiolgical conditions. Such enzymes permit the synthesis of complex metabolites like cyclic peptides in single pot sequential reactions starting from the direct precursor amino acids. In pharmaceutical research recent examples of bioengineering include the in vitro synthesis of immunomod-ulators of the cyclosporin-type and antihelminthics of the ennatin-type, as well as the in vivo synthesis of antifungals of the R106-type. The reasons for biocatalyst-assisted production are amongst others (1) the convenient and rapid access to small amounts of analogs, (2) the option to develop a later bioproduction of a certain analog since natural systems are exploited, and (3) the contribution of knowledge on these systems with a view to developing new systems for the generation of compound libraries.
- Published
- 1997
- Full Text
- View/download PDF
44. Biosynthesis of acylpeptidolactones of the daptomycin type. A comparative analysis of peptide synthetases forming A21978C and A54145
- Author
-
Horst Kleinkauf, Petra Wessels, and Hans von Döhren
- Subjects
chemistry.chemical_classification ,Sarcosine ,Streptomyces roseosporus ,biology ,Stereochemistry ,Lipoproteins ,Peptide ,Streptomyces fradiae ,Cross Reactions ,biology.organism_classification ,Biochemistry ,Streptomyces ,Amino acid ,Anti-Bacterial Agents ,Molecular Weight ,chemistry.chemical_compound ,chemistry ,Biosynthesis ,Intercellular Signaling Peptides and Proteins ,Amino Acid Sequence ,Isoleucine ,Peptide Synthases ,Peptides ,Peptide sequence - Abstract
A21978C and A54145 are antibacterial 13-residue peptides with a medium-chain-acylated amino terminus and a 10-residue lactone ring; they are produced by strains of Streptomyces roseosporus and Streptomyces fradiae, respectively. The structural differences in their peptide chains, which include amino acid replacements and modifications (L-Glu2--L-Asn, L-Asn(OH)3--L-Asp, Sar5--Gly, L-Ala6--L-Orn, L-Lys8--D-Ala, L-Asp(OMe)9--L-Asp, L-Asn11--D-Ser, and L-lle13--L-Kyn; Sar = sarcosine; L-Orn = L-ornithine, L-Kyn = L-kynurenine), reside in the multienzymatic templates directing their biosynthesis. We have examined the peptide synthetases employing immunodetection and substrate activation detected by the amino-acid-dependent ATP-PP1-exchange reaction. Two different antibodies specific for actinomycin synthetase 2 and a peptide sequence characteristic of acyl-CoA-synthetases/peptide synthetases were applied. For the A21978 system two peptide synthetases of 670 and 240 kDa were detected, together with two similar proteins of 630 and 440 kDa occurring in varying amounts. The latter are suggested to be degradation products of an unstable multienzyme. Activation of L-Asp, L-Thr, Gly, L-Orn, L-Ala and L-Ser were assigned to the high-molecular-mass components of 670, 630 and 440 kDa. The 240-kDa protein was purified to homogeneity and shown to catalyse activation of L-kynurenine. The A54145 system consisted of three peptide synthetases of 690, 590 and 250 kDa. Activations of L-Asn. L-Thr and Gly were found. The 250-kDa synthetase was capable of activating isoleucine and valine. Both systems thus show a comparable organisation; implications for the modular construction of their peptide synthetases are discussed.
- Published
- 1996
45. A nonribosomal system of peptide biosynthesis
- Author
-
Horst Kleinkauf and Hans von Döhren
- Subjects
Peptide Biosynthesis ,Molecular Sequence Data ,Sequence alignment ,Peptide ,Biology ,Biochemistry ,Methylation ,Peptides, Cyclic ,Polyketide ,chemistry.chemical_compound ,Lactones ,Structure-Activity Relationship ,Biosynthesis ,Multienzyme Complexes ,Coenzyme A Ligases ,Animals ,Aminoacylation ,Amino Acid Sequence ,Peptide sequence ,chemistry.chemical_classification ,Sequence Homology, Amino Acid ,Biological Evolution ,Amino acid ,chemistry ,Nucleic acid ,Mutagenesis, Site-Directed ,Apoproteins ,Ribosomes ,Sequence Alignment - Abstract
This review covers peptide structures originating from the concerted action of enzyme systems without the direct participation of nucleic acids. Biosynthesis proceeds by formation of linear peptidyl intermediates which may be enzymatically modified as well as transformed into specific cyclic structures. The respective enzyme systems are constructed of biosynthetic modules integrated into multienzyme structures. Genetic and DNA-sequence analysis of biosynthetic gene clusters have revealed extensive similarities between prokaryotic and eukaryotic systems, conserved principles of organisation, and a unique mechanism of transport of intermediates during elongation and modification steps involving 4'-phospho-pantetheine. These similarities permit the identification of peptide synthetases and related aminoacyl-ligases and acyl-ligases from sequence data. Similarities to other biosynthetic systems involved in the assembly of polyketide metabolites are discussed.
- Published
- 1996
46. Expression of an active adenylate-forming domain of peptide synthetases corresponding to acyl-CoA-synthetases
- Author
-
Yoen-Ok Lee, Henk van Liempt, Horst Kleinkauf, Hans von Döhren, and Ralf Dieckmann
- Subjects
Pantetheine ,Stereochemistry ,Phenylalanine ,Molecular Sequence Data ,Biophysics ,Adenylate kinase ,Biology ,Acyl-CoA-synthetase ,medicine.disease_cause ,Biochemistry ,Peptide synthetase ,Phosphates ,chemistry.chemical_compound ,Adenosine Triphosphate ,Tyrocidine synthetase ,Structural Biology ,Tyrocidine ,Panthetheine ,Coenzyme A Ligases ,Genetics ,medicine ,Escherichia coli ,Amino Acid Sequence ,Binding site ,Amino Acids ,Peptide Synthases ,Molecular Biology ,Peptide sequence ,Sequence Deletion ,chemistry.chemical_classification ,Cofactor binding ,Binding Sites ,Sequence Homology, Amino Acid ,CoA ,Cell Biology ,Adenosine Monophosphate ,Recombinant Proteins ,Amino acid ,chemistry ,Multienzyme - Abstract
Peptide synthetases and acyl-CoA-synthetases form acyl adenylates which are transferred to CoA or enzyme-bound pantetheine. To verify the existence of an adenylate domain in peptide synthetases, a 60.8 kDa fragment of tyrocidine 1-synthetase was constructed by a 1629 bp deletion, expressed in Escherichia coli, and characterized. The truncated multienzyme activated phenylalanine and substrate analogues with comparable kinetics as the over-expressed synthetase, as judged by ATP-[32P]PPi exchange reaction. Thus the N-terminal domain resembling an acyl-CoA-synthetase is an autonomous structural element. This N-terminal domain is followed by a cofactor binding domain, resembling acyl carrier proteins involved in polyketide formation.
- Published
- 1995
47. The nonribosomal peptide biosynthetic system--on the origins of structural diversity of peptides, cyclopeptides and related compounds
- Author
-
Hans von Döhren and Horst Kleinkauf
- Subjects
Peptide Biosynthesis ,Lactams ,Metabolite ,Molecular Sequence Data ,Siderophores ,Context (language use) ,Bacillus ,Biology ,Microbiology ,Peptides, Cyclic ,chemistry.chemical_compound ,Biosynthesis ,Nonribosomal peptide ,Gene cluster ,Protein biosynthesis ,Amino Acid Sequence ,Secondary metabolism ,Molecular Biology ,chemistry.chemical_classification ,Sequence Homology, Amino Acid ,Gramicidin ,Penicillium ,General Medicine ,Biological Evolution ,Cyclic peptide ,Anti-Bacterial Agents ,Biochemistry ,chemistry ,Peptides - Abstract
A variety of peptides have been detected in microorganisms. Some have found applications in various fields, for example the classical beta-lactam antibiotics, immunosuppressors like cyclosporin, promising new antibacterials like teichoplanin or daptomycin and antifungals like echinocandin. For none of these has it been established how their complicated biosynthetic pathways have evolved or what functions they fulfill within or for their producers. So it is unclear what selection processes limit the range of their structural analogues within various groups of microorganisms. We here consider recent data in the field of biosynthesis and how they may suggest mechanisms of genetic diversity. These may illustrate the complexity of genetic and intracellular organization of biosynthetic pathways and indicate the cellular context of some metabolites related to the complex background of the production of each metabolite. Research focusing on various targets like the increase of productivity of fermentations or the spread of resistances to antibacterials is slowly being understood.
- Published
- 1995
48. Identification of the ATP binding site in tyrocidine synthetase 1 by selective modification with fluorescein 5'-isothiocyanate
- Author
-
Horst Kleinkauf, W Schröder, H von Döhren, Maja Pavela-Vrančić, and Eva Pfeifer
- Subjects
Stereochemistry ,Molecular Sequence Data ,Phenylalanine ,Peptide ,Bacillus ,Biochemistry ,Peptide Mapping ,chemistry.chemical_compound ,Adenosine Triphosphate ,Nucleotide ,Amino Acid Sequence ,Fluorescein ,Binding site ,Peptide Synthases ,Molecular Biology ,chemistry.chemical_classification ,Affinity labeling ,Binding Sites ,Cell Biology ,Kinetics ,Enzyme ,chemistry ,Isothiocyanate ,Sequence Alignment ,Fluorescein-5-isothiocyanate - Abstract
Identification of the nucleotide binding site in peptide synthetases has been approached by affinity labeling of tyrocidine synthetase 1 with fluorescein 5'-isothiocyanate. Binding was accompanied by irreversible inhibition of the ATP-dependent phenylalanine activation reaction and was prevented in the presence of MgATP2-. The reaction obeyed pseudo first-order rate kinetics and was accelerated by Mg2+. Complete inhibition corresponded to incorporation of 2.3 mol of fluorescein 5'-isothiocyanate (FITC)/mol of protein. Upon protection by MgATP2-, about 1 mol of FITC is still incorporated; however, this does not affect activity. The modified synthetase was extensively fragmented by tryptic digestion and the labeled fragments isolated by reverse-phase high performance liquid chromatography. Two peptides, DHQVKIR and LDKMPLTPNDKIDR, have been identified by sequencing, and the FITC conjugate of the former peptide has been detected by laser desorption mass spectrometry. The labeled residues, Lys-422 and Lys-505, are located within highly conserved segments of this new class of synthetases.
- Published
- 1994
49. Delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase, the multienzyme integrating the four primary reactions in beta-lactam biosynthesis, as a model peptide synthetase
- Author
-
Yair Aharonowitz, Jürgen Bergmeyer, Jesus M. Cantoral, Gerald Cohen, Arnold L. Demain, Uwe Fink, Jim Kinghorn, Horst Kleinkauf, Andrew MacCabe, Harriet Palissa, Eva Pfeifer, Torsten Schwecke, Henk van Liempt, Hans von Döhren, Saul Wolfe, and Jinyou Zhang
- Subjects
ACV synthetase ,Molecular Sequence Data ,Biomedical Engineering ,Bioengineering ,Peptide ,Multifunctional Enzymes ,Tripeptide ,Biology ,beta-Lactams ,Applied Microbiology and Biotechnology ,Catalysis ,chemistry.chemical_compound ,Biosynthesis ,Valine ,Peptide synthesis ,Peptide bond ,Amino Acid Sequence ,Peptide Synthases ,chemistry.chemical_classification ,Sequence Homology, Amino Acid ,Anti-Bacterial Agents ,chemistry ,Biochemistry ,Models, Chemical ,biology.protein ,Molecular Medicine ,Sequence Alignment ,Biotechnology - Abstract
ACV synthetase forms the tripeptide precursor of penicillins and cephalosporins from alpha-aminoadipate, cysteine, and valine. Catalytic sites for substrate carboxyl activation as adenylates, peptide bond formations, epimerization and release of the tripeptide-thioester are integrated in multifunctional enzymes of 405 to 425 kD. These have been characterized from several pro- and eukaryotic beta-lactam producers. Implications of these results for the thio-template mechanism of peptide formation are discussed, as well as the use of this multienzyme as a model system for enzymatic peptide synthesis.
- Published
- 1993
50. Enzymatische Synthese biologisch aktiver Antibiotikapeptide und immunologisch suppressiver Cyclosporinderivate
- Author
-
Horst Kleinkauf
- Abstract
In der Zelle gibt es verschiedene Peptid-Biosynthesewege. Am bekanntesten ist das ribosomale System, ein komplexer Mechanismus, in dem bis zu 180 Komponenten an der Peptidbildung beteiligt sind. Mit dem ribosomalen System lassen sich lineare Proteine jeder Sequenz und Lange aus den in der Zelle fur die Proteinsynthese zur Verfugung stehenden zwanzig proteinogenen Aminosauren synthetisieren. Das Resultat ist abhangig von der jeweiligen Botenribonukleinsaure (mRNS), die vom Ribosom abgelesen wird. Das ist etwa zu vergleichen mit einem Auswahlprogramm, das auf einem Fliesband eingestellt werden kann. Die Aminosauren werden unter Adenosintriphosphat-(ATP)-verbrauch als Ami-noacyladenylate aktiviert und dann an Transfer-Ribonukleinsauren (tRNA’s) gebunden und von dort zum Ribosom transportiert. Der Einbau der verschiedenen Aminosauren in die Polypeptidkette erfolgt mit hoher Spezifitat. Der richtige Einbau wird dabei durch einen Korrekturlese-Mechanismus uberpruft. Diese Schritte sind energieaufwendig, so das pro Peptidbindung 4 bis 5,5 Hochenergie-Phosphatbindungen verbraucht werden. Die auf diese Weise synthetisierten Proteine unterliegen einem Modellierungsverfahren zum Endprodukt, wobei Aminosauresequenzen ab-oder herausgeschnitten oder auch modifiziert werden.
- Published
- 1993
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.