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Inhibitory properties of (methylene cyclopropyl) formyl-CoA (MCPF-CoA), (methylenecyclopropyl) glycine against enoyl-CoA hydratase (ECH) were studied. This helps in selection of inhibitors to control fatty acid metabolism.

Heating the dodecyl sulphate layered double hydroxide (LDH) powders in C60 containing organic solution leads to intercalation of C60 molecules in the interlayer of the double hydroxide. In the interlayer dodecyl phase, the C60 molecules exist in the dissolved state. The C60 molecules have restricted rotation compared to the free rotation in the pure solid form. The molecules experience dipolar interactions with the interlayer proton-containing species or the layer lattice atoms.

The functional characterization of a novel protein NosA in nosiheptide biosynthesis has provided an unusual C-terminal amide forming strategy in general for maturing certain amide-terminated thiopeptides by processing their precursor peptides featuring a serine extension. NosA has acted as an intermediate bearing a bis-dehydroalanine tail and has catalyzed an enamide dealkylation to remove the acrylate unit originating from the extended serine residue.

The first mechanistic insight into 2-thiosugar production in an angucycline-type antibiotic, BE-7585A, is reported. The results indicated that the sulfur insertion mechanism in 2-thiosugar biosynthesis shares similarities with that for thiamin biosynthesis.

Several flavin mononucleotide (FMN) analogues with altered electronic properties are chemoenzymatically prepared and their effects on the kinetic properties of the type II isopentenyl diphosphate:dimethylallyl diphosphate isomerase (IDI-2) catalyzed reaction are examined in order to understand the role of the reduced flavin in IDI-2 catalysis. It is shown that the reduced FMN coenzyme IDI-2 has functioned as an acid/base catalyst, with the N5 atom of the flavin playing a vital role in the deprotonation of isopentenyl pyrophosphate (IPP) en route to dimethylallyl pyrophosphate (DMAPP) formation.

Structural analysis and genetic and biochemical experiments were performed for BE-7585A produced by Amycolatopsis orientalis subsp. vinearia BA-07585 which has a 2-thiosugar and is a member of the angucycline class of compounds. The spectroscopic analyses verified the structure of BE-7585A to be closely related to rhodonocardin A and provided insight into the biosynthetic pathway for BE-7585A which could be applied for future biochemical studies of 2-thiosugar biosynthesis and BE-7585A assembly.

The catalytic properties of the spnH, spnI and spnK gene products are studied and their roles in the permethylation process of spinosyn A are validated in order to verify the functions of the rhamnose methyltransferases and also their coordination to achieve the desired level of methylation of rhamnose. The studies have shown that the completion of rhamnose permethylation is achieved by the proper control of the expression levels of the methyltransferase genes involved.

DesII from Streptomyces venezuelae is shown to consume SAM (S-adenosyl-L-methionine) versus TDP-sugar with various stoichiometries in the deamination and dehydrogenation reaction by using [Na.sub.2][S.sub.2][O.sub.4] as the reactant. The studies have not provided any evidence of an uncoupled or abortive process in the deamination reaction, as shown by the observation that dehydrogenation can take place in the absence of an external source of reductant whereas deamination cannot.

The results of the isolation and structural elucidation of a major turnover product from the enzymatic incubation of (methylenecyclopropyl)acetyl-Coa (MCPA-CoA) is presented. The mechanistic implications of the inactivation of general acyl-CoA (GAD) by MCPA-CoA is also discussed. Results indicate that the inactivation of GAD by MCPA-CoA proceeds by a radical mechanism. This suggests that GAD is capable of mediating one-electron oxidation-reduction.

Isotope-tracer experiments using deuterium-labeled amino acids have shown that aspartic acid is the precursor of the aziridine moiety and subsequent cloning and sequencing efforts have led to the identification of the azicemicin (azic) gene cluster. A possible pathway is proposed for aziridine ring formation in the azecimicins.

A strategy to identify the modification sites on multimodular domain D [poly(ADP-ribose) polymerase-1 (PARP-1)] using the PARP-1 E988Q mutant, which only catalyzes mono(ADP-ribosyl)ation is presented. Two synthetic peptides with amino acid sequences derived from the ADP-ribosylation sites of domain D that act as PARP-1 substrates which could be applied for future studies of PARP-1 catalysis is also demonstrated.

The purification and reconstitution of [His.sub.6]-tagged DesII, the characterization of its [4Fe-4S] cluster and the capability of flavodoxin, flavodoxin reductase and NADPH to reduce the [[4Fe-4S].sup.2+] cluster are described. The analysis of deuterium incorporation into S-adenosyl-L-methionine (SAM) using TDP-[3-[super 2]H]-4-amino-4,6-dideoxy-D-glucose as the substrate has provided evidence for direct hydrogen atom transfer to a 5'-deoxyadenosyl radical in the catalytic cycle.

The various mechanisms involved in the copolymerization of [CO.sub.2] with epoxides via a thermally stable single-site cobalt(III) catalyst are explained. The cobalt-based catalyst is shown to exhibit brilliant activity and selectivity, hence enhancing the process of polymer formation.

The secondary kinetic isotope effects (KIEs) measured in the equilibrium perturbation experiments with [3-[super 2]H]- and [4-[super 2]H]-1-deoxy-D-xylulose 5-phosphate (DXP) has supported a retroaldol/aldol mechanism for the DXP reductoisomerase (DXR) catalyzed rearrangement of DXP to methyl-D-erythritol 4-phosphate (MEP). The mechanistic information will help in the future design of specific inhibitors for DXR, which are in turn used for developing new antibacterial agents.

The [super 18]O kinetic isotope effects (KIEs) of different non-heme iron enzymes with different reductants are calculated to explain the [O.sub.2] activation process taking place in them. These enzymes are shown to be widely used in the catalysis of a large number of oxygenation and oxidation reactions.

D-Forosamine biosynthesis is studied by using the five genes from the spinosyn gene cluster involved in the conversion ofTDP-4-keto-6-deoxy-D-glucose to TDP-D-forosamine. The steady-state kinetic parameters for the SpnQ-catalyzed reaction and the substrate specificities of SpnQ and SpnR are explored.

The biosynthesis of spinosyn macrolides in Saccharopolyspora spinosa is discussed. The analysis reveals that these macrolides act as the cross-linking precursors with SpnJ acting as the catalyst.

The characterization of the enediyne antitumor antibiotic maduropeptin (MDP) gene cluster obtained from Actinomadura madurae ATCC 39144 is studied. The results have proved to be of great help for the biosynthesis of several enediyne polyketides.

An engineered biosynthetic method is introduced that produces non-natural sugar-bearing macrolide derivatives in vivo. The preparation of nine non-natural sugar-bearing compounds is described and the feasibility of the technique to develop non-natural sugar-bearing secondary metabolites is discussed.

The biosynthesis of D-forosamine that was investigated with a view to characterize unusual sugar biosynthetic pathways and to find genes that are useful for in vivo glycodiversification of secondary metabolites is discussed. The results illustrate the role of SpnQ in the biosynthesis of D-forosamine, the catalyzed reaction of which is very different from that of the COID reaction.

The aglycone is biosynthesized by the sequential condensation of acyl thioesters catalyzed polyketide synthases (PKS). The results show that the linear polyketide chain 8 can serve as a chemically competent substrate for macrolide glycosyltransferase discovery of an auxiliary protein (DesVII).

Sequence analysis of genes found in the biosynthetic gene clusters has resulted in tentative assignments for the encoded proteins and their putative functions in the biosynthetic pathways. Critical evidence assigning the roles of the key enzymes involved in the TDP-L-mycarose biosynthesis and complete the reconstitution of the entire mycarose pathway in vitro is provided.

The in vitro activity of a macrolide glycosyltransferases (GT), DesVII is demonstrated which catalyzes the attachment of TDP-D-desosamine to either the 12- or the 14-membered macrolactone ring to make methymycin/neomethymycin and narbomycin/pikro-mycin in Streptomyces venezuelae. The enzyme is very dependent on an auxilliary protein, DesVIII, for activity.

Resonance Raman spectroscopic techniques and quinone staining are used to identify and study the biogenesis of hydroypropylphosphonic acid epoxidase (HppE). It is found that HppE can act as an oxygenase through a putative high valent iron-oxo or an iron-hydroperoxo intermediate.

It is proposed that altering the electrochemical properties of the reaction center can change regiospecificity of hydride transfer catalyzed by a pyridine nucleotide-dependent enzyme. The difluoromethylene functionality has assumed a role as a carbonyl mimic with an apparently reversed regioselectivity of hydride reduction.

Several experiments are conducted to reveal a new two-stage one-pot method for the enzymatic biosynthesis of TDP-L-mycarose from the readily available precursors, glucose-1-phosphate and thymidine. The process described demonstrates the use of enzymes in the synthesis of several unstable TDP-sugar derivatives and hence can be widely used for the identification of more such enzymes in addition to the synthesis of other unusual sugars.

The catalytic role of Tylla as TDP-4-keto-6-deoxy-D-glucose 3,4-isomerase is firmly established. The finding is significant as it assigns a function to an orphan open reading frames (ORF) whose encoded protein catalyzes an important conversion in unusual sugar biosynthesis.

The delineation of the biosynthesis of the (S)-3-chloro-4,5-dihydroxy-beta-phenylalanine moiety from L-tyrosine and its incorporation into the C-1027 chromophore, which are catalyzed by six enzymes, by in vivo and in vitro studies of five of the six enzymes are reported. A novel strategy for beta-amino acid modification is provided, featuring beta-amino acyl-S-carrier protein intermediates, and also, shed light into engineering the C-1027 biosynthetic machinery for the production of novel analogues as exemplified by 20-deschloro-C-1027 and 20-deschloro-22-deshydroxy-C-1027.

The biosynthesis of D-forosamine, 4-N,N-(dimethylamino)-2,3,4,6-tetradeoxy-alpha-D-threo-hexopyranose is investigated to characterize deoxysugar biosynthetic pathways and to collect genes useful for in vivo glycodiversification of secondary metabolites. The results obtained provide the first evidence establishing the relevance of the spnR gene to forosamine biosynthesis.

The Strptomyces venezuelae strains containing selected genes from the L-dihydrostreptose pathway are constructed and the biosynthetic implications obtained for the dihydrostreptose pathway are reported. The versatility of the glycosyltransferase on substrate selection highlights its potential as a catalyst in the construction of new macrolides carrying a broad range of modified sugars.

Research is presented demonstrating the direct association of a radical with the PMP coenzyme in E1 catalysis, using EPR and isotopic PMP labeling techniques.

The rechargeable lithium-sulfur battery is a promising option for energy storage applications because of its low cost and high energy density. The electrochemical performance of the sulfur cathode, however, is substantially compromised because of fast capacity decay caused by polysulfide dissolution/shuttling and low specific capacity caused by the poor electrical conductivities of the active materials. Herein we demonstrate a novel strategy to address these two problems by designing and synthesizing a carbon nanotube (CNT)/NiFe2O4-S ternary hybrid material structure. In this unique material architecture, each component synergistically serves a specific purpose: The porous CNT network provides fast electron conduction paths and structural stability. The NiFe2O4 nanosheets afford strong binding sites for trapping polysulfide intermediates. The fine S nanoparticles well-distributed on the CNT/NiFe2O4 scaffold facilitate fast Li+ storage and release for energy delivery. The hybrid material exhibits balanced high performance with respect to specific capacity, rate capability, and cycling stability with outstandingly high Coulombic efficiency. Reversible specific capacities of 1350 and 900 mAh g-1 are achieved at rates of 0.1 and 1 C respectively, together with an unprecedented cycling stability of ~0.009% capacity decay per cycle over more than 500 cycles. [ABSTRACT FROM AUTHOR]

EPR studies on the pyridoxamine 5'-phosphate (PMP) induced catalysis of the PMP-dependent enzyme, CDP-6-deoxy-L-threo-D-glycero-4-hexulose-3-dehydrase (E1), reveals that PMP mediates anion induced dehydration and is an E3-electron receptor in the redox reaction. The presence of the E1 (2Fe-2S) center and the radical intermediate proves that the PMP-induced enzyme redox reactions have a radical mechanism. PMP is the amine form of the B6 coenzyme.

The gene coding for E1(ascC) from a Yersinia pseudotuberculosis lambda gt11 subgenomic library was cloned through hybridization with a degenerate oligonucleotide probe built from the N-terminal sequence of the purified wild-type E. This was done to investigate the biosynthesis of 3, 6-dideoxyhexoses, and represents the characterization of the first pyridoxamine 5'-phosphate (PMP)-dependent iron-sulfur containing enzyme needed for this biosynthesis.