Molecular and biochemical studies of the Bacillus subtilis FtsY protein, a homologue of the α subunit of mammalian signal recognition particle

In mammalian cellls, the signal recognition particle(SRP) and the SRP receptor play a central role in targeting presecretory proteins to the membrane of the endoplasmic reticulum(ER). SRP is a ribonucleo-protein complex composed of one RNA(SRP 7S RNA) molecule and six proteins of 9, 14, 19, 54, 68, and 72 kDa. SRP interacts with the signal sequence of nascent polypeptide emerging from ribosomes and the complex formed is targeted and bound to a hetero-dimeric receptor consisting of SRα and SRβ on the ER. In contrast, it has been considered that the chaperones and Sec proteins play a pivotal role in targeting and translocation of secretory proteins in Escherichia coli. However, recently, the molecular homologues to the components of mammalian SRP have been identified in prokaryotes. In E. coli, 4.5S RNA and E. coli Ffh protein have been identified as homologues to the SRP 7S RNA and SRP54 of mammalian SRP, respectively. Furthermore, FtsY exhibits a homology with SRα. In Bacillus subtilis, small cytoplasmic RNA(scRNA) and B. subtilis Ffh protein have been also identified as homologues to the SRP 7S RNA and SRP54, respectively. Depletion of either the SRP RNA or SRP54 homologues ･･･Thesis (Ph. D. in Sciences)--University of Tsukuba, (B), no. 1714, 2001.3.23Includes bibliographical referencesTitlepage,Contens -- Abbreviations,Abstracts -- General Introduction -- Section I. Cloning and Characterization of the Bacillus subtilis ftsY (srb) Which product is a Homologue of the α-Subunit of Mammalian Signal Recognition Particle Receptor -- Section II. Expression of the ftsY Gene is Controlled by Different Promoters in Vegetative and Sporulating Cells of Bacillus subtilis, and functional analysis of FtsY protein -- General Discussion -- References -- Acknowledgement

Enhanced Recombinant Protein Productivity by Genome Reduction in Bacillus subtilis

The emerging field of synthetic genomics is expected to facilitate the generation of microorganisms with the potential to achieve a sustainable society. One approach towards this goal is the reduction of microbial genomes by rationally designed deletions to create simplified cells with predictable behavior that act as a platform to build in various genetic systems for specific purposes. We report a novel Bacillus subtilis strain, MBG874, depleted of 874 kb (20%) of the genomic sequence. When compared with wild-type cells, the regulatory network of gene expression of the mutant strain is reorganized after entry into the transition state due to the synergistic effect of multiple deletions, and productivity of extracellular cellulase and protease from transformed plasmids harboring the corresponding genes is remarkably enhanced. To our knowledge, this is the first report demonstrating that genome reduction actually contributes to the creation of bacterial cells with a practical application in industry. Further systematic analysis of changes in the transcriptional regulatory network of MGB874 cells in relation to protein productivity should facilitate the generation of improved B. subtilis cells as hosts of industrial protein production

MOESM2 of A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

Additional file 2: Figure S2. SDS-PAGE of enzyme preparation JN11H and JNK25H. SDS-PAGE was carried out with Any kD Mini-PROTEAN TGX Precast Protein Gels (Bio-Rad, Hercules, CA) and the gel was activated and imaged using the ChemiDoc MP imaging system (Bio-Rad).3 μg of protein were loaded on each gel. Lanes: M, Precision Plus Protein Unstained Standard; 1, JN11H enzyme preparation produced by T. reesei strain X3AB1; 2, JNK25H enzyme preparation produced by strain X2PX10. Cellobiohydrolases (CBH 1, CBH2), endoglucanases (EG1 and EG2), xylanases (XYN1 and XYN2) and β-xylosidases (BXL) from Trichoderma reesei, and heterologously expressed proteins (β-glucosidase AaBGL1 from Aspergillus aculeatus and xylanase PspXyn10 from Penicillium sp. KSM-F532) are shown

MOESM1 of A novel GH10 xylanase from Penicillium sp. accelerates saccharification of alkaline-pretreated bagasse by an enzyme from recombinant Trichoderma reesei expressing Aspergillus β-glucosidase

Additional file 1: Figure S1. Multiple alignments and phygogenetic tree of PspXyn10, PspXyn10 orthologs and TrXyn3. (a) Multiple alignments of the PspXyn10, PspXyn10 orthologs and TrXyn3. The amino acid sequence of PspXyn10 was compared with those of endo-1,4-beta-xylanase from Penicillium brasilianum (OOQ87260), endo-1,4-beta-xylanase D from Penicillium subrubescens (OKO90312), endo-1,4-beta-xylanase from Rasamsonia emersonii CBS 393.64 (XP_013330999), hypothetical protein PENANT_c053G02808 from Penicillium antarcticum (OQD79351), endo-1,4-beta-xylanase Xyl10A from Talaromyces cellulolyticus (GAM37231) and TrXyn3 from Trichoderma reesei (BAA89465) by multiple alignment. The alignment was created using ClustalW2 on Genetyx Version 12 software (Genetyx). Amino acid numbers are shown on the left and right. Black boxes indicate invariant residues. Gray boxes indicate the residues conserved in more than half of aligned sequences. (b) Phylogenetic tree of the PspXyn10, PspXyn10 orthologs and TrXyn3. Phylogenetic tree of the amino acid sequences were created using the NJ method under 1000 times bootstrap conditions using Genetyx Version 12 software (Genetyx)

Mannitol-1-Phosphate Dehydrogenase (MtlD) Is Required for Mannitol and Glucitol Assimilation in Bacillus subtilis: Possible Cooperation of mtl and gut Operons

We found that mannitol-1-phosphate dehydrogenase (MtlD), a component of the mannitol-specific phosphotransferase system, is required for glucitol assimilation in addition to GutR, GutB, and GutP in Bacillus subtilis. Northern hybridization of total RNA and microarray studies of RNA from cells cultured on glucose, mannitol, and glucitol indicated that mannitol as the sole carbon source induced hyperexpression of the mtl operon, whereas glucitol induced both mtl and gut operons. The B. subtilis mtl operon consists of mtlA (encoding enzyme IICBA(mt1)) and mtlD, and its transcriptional regulator gene, mtlR, is located 14.4 kb downstream from the mtl operon on the chromosome. The mtlA, mtlD, and mtlR mutants disrupted by the introduction of the pMUTin derivatives MTLAd, MTLDd, and MTLRd, respectively, could not grow normally on either mannitol or glucitol. However, the growth of MTLAd on glucitol was enhanced by IPTG (isopropyl-β-d-thiogalactopyranoside). This mutant has an IPTG-inducible promoter (Pspac promoter) located in mtlA, and this site corresponds to the upstream region of mtlD. Insertion mutants of mtlD harboring the chloramphenicol resistance gene also could not grow on either mannitol or glucitol. In contrast, an insertion mutant of mtlA could grow on glucitol but not on mannitol in the presence or absence of IPTG. MtlR bound to the promoter region of the mtl operon but not to a DNA fragment containing the gut promoter region