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Mirror-image proteins, composed of d-amino acids, are an attractive therapeutic modality, as they exhibit high metabolic stability and lack immunogenicity. Development of mirror-image binding proteins is achieved through chemical synthesis of d-target proteins, phage display library selection of l-binders and chemical synthesis of (mirror-image) d-binders that consequently bind the physiological l-targets. Monobodies are well-established synthetic (l-)binding proteins and their small size (~90 residues) and lack of endogenous cysteine residues make them particularly accessible to chemical synthesis. Here, we develop monobodies with nanomolar binding affinities against the d-SH2 domain of the leukemic tyrosine kinase BCR::ABL1. Two crystal structures of heterochiral monobody-SH2 complexes reveal targeting of the pY binding pocket by an unconventional binding mode. We then prepare potent d-monobodies by either ligating two chemically synthesized d-peptides or by self-assembly without ligation. Their proper folding and stability are determined and high-affinity binding to the l-target is shown. d-monobodies are protease-resistant, show long-term plasma stability, inhibit BCR::ABL1 kinase activity and bind BCR::ABL1 in cell lysates and permeabilized cells. Hence, we demonstrate that functional d-monobodies can be developed readily. Our work represents an important step towards possible future therapeutic use of d-monobodies when combined with emerging methods to enable cytoplasmic delivery of monobodies. In this work, the authors develop mirror-image monobodies (Mb) made of D-amino acids against the BCR::ABL1 SH2 domain with high binding affinities. The heterochiral Mb-SH2 structures reveal an unusual binding mode. The D-Mbs are protease-resistant, inhibit BCR::ABL1 kinase activity and bind BCR::ABL1 in cell lysates. [ABSTRACT FROM AUTHOR]

Background: This clinical trial explores the Sigstad score for late dumping syndrome in postoperative patients who have undergone sleeve gastrectomy (SG) or One Anastomosis Gastric Bypass (OAGB). The aims of this study are to investigate the correlations with late dumping syndrome, to evaluate the reliability and validity of the Sigstad score and to discuss a modified scoring system. Methods: The study was conducted at the Obesity Center of the Westküstenklinikum Heide and included 271 patients. Data collection involved conducting interviews, diet diaries and measuring blood glucose levels. Non-parametric tests, logistic regression and McDonald's Omega were the selected statistical approaches. Results: Body Mass Index (BMI) decreased over time (-9.67 kg/m2 at 4 months, -15.58 kg/m2 at 12 months). Preoperatively, the Sigstad score exhibited the highest value, and no occurrences of late dumping syndrome were observed. No significant differences were found in BMI concerning late dumping syndrome or Sigstad score among postoperative patients. Postoperative patients experienced an increase in gastrointestinal symptoms. The reliability test showed a McDonald's omega value of 0.509. The analysis conducted through binary logistic regression indicated dizziness as a significant predictor of late dumping syndrome; however, this finding did not hold up after performing Bonferroni correction. Conclusion: The Sigstad score is not a reliable or valid method for detecting late dumping syndrome after surgery for obesity and metabolic disorders. It is necessary to have alternatives that use objective measures and assess the quality of life, and that these alternatives be validated in large patient cohorts. [ABSTRACT FROM AUTHOR]

Inhibition of eukaryotic initiation factor 4A has been proposed as a strategy to fight pathogens. Rocaglates exhibit the highest specificities among eIF4A inhibitors, but their anti-pathogenic potential has not been comprehensively assessed across eukaryotes. In silico analysis of the substitution patterns of six eIF4A1 aa residues critical to rocaglate binding, uncovered 35 variants. Molecular docking of eIF4A:RNA:rocaglate complexes, and in vitro thermal shift assays with select recombinantly expressed eIF4A variants, revealed that sensitivity correlated with low inferred binding energies and high melting temperature shifts. In vitro testing with silvestrol validated predicted resistance in Caenorhabditiselegans and Leishmaniaamazonensis and predicted sensitivity in Aedes sp., Schistosomamansoni, Trypanosomabrucei, Plasmodiumfalciparum, and Toxoplasmagondii. Our analysis further revealed the possibility of targeting important insect, plant, animal, and human pathogens with rocaglates. Finally, our findings might help design novel synthetic rocaglate derivatives or alternative eIF4A inhibitors to fight pathogens. [ABSTRACT FROM AUTHOR]

The formation of C-C bonds by using CoA independent acyltransferases may have significant impact for novel methods for biotechnology. We report the identification of Pseudomonas strains with CoA-independent acyltransferase activity as well as the heterologous expression of the enzyme in E. coli. The cloning strategies and selected expression studies are discussed. The recombinant acyltransferases were characterized with regard to thermal and storage stability, pH,- and co-solvent tolerance. Moreover, the impact of bivalent metals, inhibitors, and other additives was tested. Careful selection of expression and working conditions led to obtain recombinant acyltransferase form Pseudomonas protegens with up to 11 U mL−1 activity. [ABSTRACT FROM AUTHOR]

Main conclusion: We present a comprehensive overview on flavonoid-related phenotypes of A. thaliana tt and tds mutants, provide tools for their characterisation, increase the number of available alleles and demonstrate that tds3 is allelic to tt12 and tds5 to aha10. Flavonoid biosynthesis is one of the best-studied secondary metabolite pathways in plants. In the model system Arabidopsis thaliana it leads to the synthesis of three phenolic compound classes: flavonol glycosides, anthocyanins and proanthocyanidins (PAs). PAs appear brown in their oxidised polymeric forms, and most A. thaliana mutants impaired in flavonoid accumulation were identified through screens for lack of this seed coat pigmentation. These mutants are referred to as transparent testa ( tt) or tannin-deficient seed ( tds). More than 20 mutants of these types have been published, probably representing most of the genes relevant for PA accumulation in A. thaliana. However, data about the genes involved in PA deposition or oxidation are still rather scarce. Also, for some of the known mutants it is unclear if they represent additional loci or if they are allelic to known genes. For the present study, we have performed a systematic phenotypic characterisation of almost all available tt and tds mutants and built a collection of mutants in the genetic background of the accession Columbia to minimise effects arising from ecotype variation. We have identified a novel tt6 allele from a forward genetic screen and demonstrated that tds3 is allelic to tt12 and tds5 to aha10. [ABSTRACT FROM AUTHOR]

Objectives: To compare the accuracy of cone-beam CT (CBCT) and multislice CT (MSCT) with regard to its use in image-guided dental implant surgery in a prospective model based study. Material and methods: Ten photopolymer-acrylate mandibula models, each with four precise metal reference markers, were scanned with MSCT and CBCT. The six reference distances between the markers were measured by a three-axis milling machine first. The distances were then measured by (1) navigation with the Medtronic StealthStation® TREON™ image-guided surgery system, (2) with the Medtronic planning-tool and (3) on the PC with the Mimics® software. Mean values were calculated for all three methods for CBCT and MSCT and were compared for statistical significance. Results: Of all measurements, 83% of the arithmetic mean values were within the ±0.5 mm range (MSCT 88% and CBCT 78%) and 17% within the ±1.0 mm range (MSCT 12% and CBCT 22%). The absolute difference of the arithmetic mean values showed no statistically significant difference between MSCT and CBCT. The difference of the overall mean values to the reference was 0.43 mm for MSCT and 0.46 mm for CBCT. Conclusions: The data of our study prove that the application of CBCT for the indicated purpose yielded good results comparable to those of MSCT. All three measuring methods were feasible and accuracy was statistically not different between the data acquired by MSCT and CBCT within the setting of this study. [ABSTRACT FROM AUTHOR]