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de Jong A.,University of Groningen | van Heel A.J.,University of Groningen | Kok J.,University of Groningen | Kuipers O.P.,University of Groningen | Kuipers O.P.,Kluyver Center for Genomics of Industrial Fermentation
Nucleic Acids Research

Mining bacterial genomes for bacteriocins is a challenging task due to the substantial structure and sequence diversity, and generally small sizes, of these antimicrobial peptides. Major progress in the research of antimicrobial peptides and the ever-increasing quantities of genomic data, varying from (un)finished genomes to meta-genomic data, led us to develop the significantly improved genome mining software BAGEL2, as a follow-up of our previous BAGEL software. BAGEL2 identifies putative bacteriocins on the basis of conserved domains, physical properties and the presence of biosynthesis, transport and immunity genes in their genomic context. The software supports parameter-free, class-specific mining and has high-throughput capabilities. Besides building an expert validated bacteriocin database, we describe the development of novel Hidden Markov Models (HMMs) and the interpretation of combinations of HMMs via simple decision rules for prediction of bacteriocin (sub-)classes. Furthermore, the genetic context is automatically annotated based on (combinations of) PFAM domains and databases of known context genes. The scoring system was fine-tuned using expert knowledge on data derived from screening all bacterial genomes currently available at the NCBI. BAGEL2 is freely accessible at http://bagel2.molgenrug.nl. © The Author(s) 2010. Published by Oxford University Press. Source

Boonstra M.,University of Groningen | de Jong I.G.,University of Groningen | Scholefield G.,Northumbria University | Murray H.,Northumbria University | And 3 more authors.
Molecular Microbiology

When starved, Bacillus subtilis cells can enter the developmental programme of endospore formation by activation of the master transcriptional regulator Spo0A. Correct chromosome copy number is crucial for the production of mature and fully resistant spores. The production and maintenance of one chromosome for the mother cell and one copy for the forespore requires accurate co-ordination between DNA replication and initiation of sporulation. Here, we show that Spo0A regulates chromosome copy number by directly binding to a number of Spo0A binding sites that are present near the origin of replication (oriC). We demonstrate that cells lacking three specific Spo0A binding sites at oriC display increased chromosome copy numbers when sporulation is induced. Our data support the hypothesis that Spo0A directly controls DNA replication during sporulation by binding to oriC. © 2013 Blackwell Publishing Ltd. Source

de Jong I.G.,University of Groningen | Veening J.-W.,University of Groningen | Kuipers O.P.,Kluyver Center for Genomics of Industrial Fermentation
Environmental Microbiology

How cells dynamically respond to fluctuating environmental conditions depends on the architecture and noise of the underlying genetic circuits. Most work characterizing stress pathways in the model bacterium Bacillus subtilis has been performed on bulk cultures using ensemble assays. However, investigating the single cell response to stress is important since noise might generate significant phenotypic heterogeneity. Here, we study the stress response to carbon source starvation and compare both population and single cell data. Using a top-down approach, we investigate the transcriptional dynamics of various stress-related genes of B. subtilis in response to carbon source starvation and to increased cell density. Our data reveal that most of the tested gene-regulatory networks respond highly heterogeneously to starvation and cells show a large degree of variation in gene expression. The level of highly dynamic diversification within B. subtilis populations under changing environments reflects the necessity to study cells at the single cell level. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd. Source

Kovacs A.T.,University of Groningen | Kuipers O.P.,University of Groningen | Kuipers O.P.,Kluyver Center for Genomics of Industrial Fermentation
Journal of Bacteriology

Transcriptome analysis of a Bacillus subtilis rok strain that showed reduced complex colony structure formation revealed significant downregulation of the yuaB gene. Overexpression of yuaB restored structure formation in the rok strain. We show that transcription of yuaB is indirectly regulated by Rok, independently from its previously described AbrB-dependent regulation. Copyright © 2011, American Society for Microbiology. All Rights Reserved. Source

Khusainov R.,University of Groningen | Heils R.,University of Groningen | Lubelski J.,University of Groningen | Moll G.N.,LanthioPep | And 2 more authors.
Molecular Microbiology

Although nisin is a model lantibiotic, our knowledge of the specific interactions of prenisin with its modification enzymes remains fragmentary. Here, we demonstrate that the nisin modification enzymes NisB and NisC can be pulled down in vitro from Lactococcus lactis by an engineered His-tagged prenisin. This approach enables us to determine important intermolecular interactions of prenisin with its modification machinery within L.lactis. We demonstrate that (i) NisB has stronger interactions with precursor nisin than NisC has, (ii) deletion of the propeptide part keeping the nisin leader intact leads to a lack of binding, (iii) NisB point mutants of highly conserved residues W616, F342A, Y346F and P639A are still able to dehydrate prenisin, (iv) NisB Δ(77-79)Y80F mutant decreased the levels of NisB-prenisin interactions and resulted in unmodified prenisin, (v) substitution of an active site residue H331A in NisC leads to higher amounts of the co-purified complex, (vi) NisB is present in the form of a dimer, and (vii) the region FNLD (-18 to -15) of the leader is an important site for binding not only to NisB, but also to NisC. © 2011 Blackwell Publishing Ltd. Source

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