The Bacillus subtilis signaling protein SpoIVB defines a new family of serine peptidases

The protein SpoIVB plays a key role in signaling in the sigma (K) checkpoint of Bacillus subtilis. This regulatory mechanism coordinates late gene expression during development in this organism and we have recently shown SpoIVB to be a serine peptidase. SpoIVB signals by transiting a membrane, undergoing self-cleavage, and then by an unknown mechanism activating a zinc metalloprotease, SpoIVFB, which cleaves pro-sigma (K) to its active form, sigma (K), in the outer mother cell chamber of the developing cell. In this work we have characterized the serine peptidase domain of SpoIVB. Alignment of SpoIVB with homologues from other spore formers has allowed site-specific mutagenesis of all potential active site residues within the peptidase domain. We have defined the putative catalytic domain of the SpoIVB serine peptidase as a 160-amino-acid residue segment at the carboxyl terminus of the protein. His236 and Ser378 are the most important residues for proteolysis, with Asp363 being the most probable third member of the catalytic triad. In addition, we have shown that mutations at residues Asn290 and His394 lead to delayed signaling in the sigma (K) checkpoint. The active site residues suggest that SpoIVB and its homologues from other spore formers are members of a new family of serine peptidases of the trypsin superfamily

Emerging Applications of Bacterial Spores in Nanobiotechnology

Bacterial spores are robust and dormant life forms with formidable resistance properties, in part, attributable to the multiple layers of protein that encase the spore in a protective and flexible shield. The coat has a number of features pertinent to the emerging field of nanobiotechnology including self-assembling protomers and the capacity for engineering and delivery of foreign molecules. This review gives an account of recent progress describing the use of the spore, and specifically, the spore coat as a vehicle for heterologous antigen presentation and protective immunization (vaccination). As interest in the spore coat increases it seems likely that they will be exploited further for drug and enzyme delivery as well as a source of novel self-assembling proteins

The PDZ domain of the SpoIVB serine peptidase facilitates multiple functions

During spore formation in Bacillus subtilis, the SpoIVB protein is a critical component of the sigma (K) regulatory checkpoint. SpoIVB has been shown to be a serine peptidase that is synthesized in the spore chamber and which self-cleaves, releasing active forms. These forms can signal proteolytic processing of the transcription factor sigma (K) in the outer mother cell chamber of the sporulating cell. This forms the basis of the sigma (K) checkpoint and ensures accurate sigma (K)-controlled gene expression. SpoIVB has also been shown to activate a second distinct process, termed the second function, which is essential for the formation of heat-resistant spores. In addition to the serine peptidase domain, SpoIVB contains a PDZ domain. We have altered a number of conserved residues in the PDZ domain by site-directed mutagenesis and assayed the sporulation phenotype and signaling properties of mutant SpoIVB proteins. Our work has revealed that the SpoIVB PDZ domain could be used for up to four distinct processes, (i) targeting of itself for trans proteolysis, (11) binding to the protease inhibitor BofC, (iii) signaling of pro-sigma (K) processing, and (iv) signaling of the second function of SpoIVB

Carbohydrate-active enzymes from pigmented Bacilli: a genomic approach to assess carbohydrate utilization and degradation

<p>Abstract</p> <p>Background</p> <p>Spore-forming <it>Bacilli </it>are Gram-positive bacteria commonly found in a variety of natural habitats, including soil, water and the gastro-intestinal (GI)-tract of animals. Isolates of various <it>Bacillus </it>species produce pigments, mostly carotenoids, with a putative protective role against UV irradiation and oxygen-reactive forms.</p> <p>Results</p> <p>We report the annotation of carbohydrate active enzymes (CAZymes) of two pigmented <it>Bacilli </it>isolated from the human GI-tract and belonging to the <it>Bacillus indicus </it>and <it>B. firmus </it>species. A high number of glycoside hydrolases (GHs) and carbohydrate binding modules (CBMs) were found in both isolates. A detailed analysis of CAZyme families, was performed and supported by growth data. Carbohydrates able to support growth as the sole carbon source negatively effected carotenoid formation in rich medium, suggesting that a catabolite repression-like mechanism controls carotenoid biosynthesis in both <it>Bacilli</it>. Experimental results on biofilm formation confirmed genomic data on the potentials of <it>B. indicus </it>HU36 to produce a levan-based biofilm, while mucin-binding and -degradation experiments supported genomic data suggesting the ability of both <it>Bacilli </it>to degrade mammalian glycans.</p> <p>Conclusions</p> <p>CAZy analyses of the genomes of the two pigmented <it>Bacilli</it>, compared to other <it>Bacillus </it>species and validated by experimental data on carbohydrate utilization, biofilm formation and mucin degradation, suggests that the two pigmented <it>Bacilli </it>are adapted to the intestinal environment and are suited to grow in and colonize the human gut.</p

Inverse folding for antibody sequence design using deep learning

We consider the problem of antibody sequence design given 3D structural information. Building on previous work, we propose a fine-tuned inverse folding model that is specifically optimised for antibody structures and outperforms generic protein models on sequence recovery and structure robustness when applied on antibodies, with notable improvement on the hypervariable CDR-H3 loop. We study the canonical conformations of complementarity-determining regions and find improved encoding of these loops into known clusters. Finally, we consider the applications of our model to drug discovery and binder design and evaluate the quality of proposed sequences using physics-based methods.Comment: 2023 ICML Workshop on Computational Biology, model weights available at https://zenodo.org/record/816469