Clostridium kluyveri is unique among the clostridia; it grows anaerobically on ethanol and acetate as sole energy sources. Fermentation products are butyrate, caproate, and H2. We report here the genome sequence of C. kluyveri, which revealed new insights into the metabolic capabilities of this well studied organism. A membrane-bound energy-converting NADH:ferredoxin oxidoreductase (RnfCDGEAB) and a cytoplasmic butyryl-CoA dehydrogenase complex (Bcd/EtfAB) coupling the reduction of crotonyl-CoA to butyryl-CoA with the reduction of ferredoxin represent a new energy-conserving module in anaerobes. The genes for NAD-dependent ethanol dehydrogenase and NAD(P)-dependent acetaldehyde dehydrogenase are located next to genes for microcompartment proteins, suggesting that the two enzymes, which are isolated together in a macromolecular complex, form a carboxysome-like structure. Unique for a strict anaerobe, C. kluyveri harbors three sets of genes predicted to encode for polyketide/nonribosomal peptide synthetase hybrides and one set for a nonribosomal peptide synthetase. The latter is predicted to catalyze the synthesis of a new siderophore, which is formed under iron-deficient growth conditions. butyryl-CoA dehydrogenase ͉ electron transfer flavoproteins ͉ genome sequence ͉ Rnf-dependent energy conservation

The genome of Bacillus licheniformis DSM13 consists of a single chromosome that has a size of 4,222,748 base pairs. The average G+C ratio is 46.2%. 4,286 open reading frames, 72 tRNA genes, 7 rRNA operons and 20 transposase genes were identified. The genome shows a marked co-linearity with Bacillus subtilis but contains defined inserted regions that can be identified at the sequence as well as at the functional level. B. licheniformis DSM13 has a well-conserved secretory system, no polyketide biosynthesis, but is able to form the lipopeptide lichenysin. From the further analysis of the genome sequence, we identified conserved regulatory DNA motives, the occurrence of the glyoxylate bypass and the presence of anaerobic ribonucleotide reductase explaining that B. licheniformis is able to grow on acetate and 2,3-butanediol as well as anaerobically on glucose. Many new genes of potential interest for biotechnological applications were found in B. licheniformis; candidates include proteases, pectate lyases, lipases and various polysaccharide degrading enzymes.

SNAREs on transport vesicles and target membranes are required for vesicle targeting and fusion. Here we describe a novel yeast protein with a typical SNARE motif but with low overall amino acid homologies to other SNAREs. The protein localized to the endoplasmic reticulum (ER) and was therefore named Use1p (unconventional SNARE in the ER). A temperature‐sensitive use1 mutant was generated. use1 mutant cells accumulated the ER forms of carboxypeptidase Y and invertase. More specific assays revealed that use1 mutant cells were defective in retrograde traffic to the ER. This was supported by strong genetic interactions between USE1 and the genes encoding SNAREs in retrograde traffic to the ER. Antibodies directed against Use1p co‐immunoprecipitated the SNAREs Ufe1p, myc‐Sec20p and Sec22p, which form a SNARE complex required for retrograde traffic from the Golgi to the ER, but neither Bos1p nor Bet1p (members of the SNARE complex in anterograde traffic to the Golgi). Therefore, we conclude that Use1p is a novel SNARE protein that functions in retrograde traffic from the Golgi to the ER.

The glucose and nitrogen starvation stimulons of Bacillus licheniformis were determined by transcriptome and proteome analyses. Under both starvation conditions, the main response of B. licheniformis was a switch to the usage of alternative nutrient sources. This was indicated by an induction of genes involved in the metabolism of C-2 substrates during glucose limitation. In addition, B. licheniformis seems to be using other organic substances like amino acids and lipids as carbon sources when subjected to glucose starvation. This observation is supported by the induction of a high number of genes coding for proteins involved in amino acid and lipid degradation. During nitrogen starvation, genes for several proteases and peptidases involved in nitrate and nitrite assimilation were induced, which enables this bacterium to recruit nitrogen from alternative sources. Both starvation conditions led to a down-regulation of transcription of most vegetative genes, which was subsequently reflected by a reduced synthesis of the corresponding proteins. A selected set of genes was induced by both starvation conditions. Among them were yvyD, citA and the putative methylcitrate shunt genes mmgD, mmgE and yqiQ. However, both starvation conditions did not induce a general SigmaB-dependent stress response.

The envelope is an essential structure of the bacterial cell, and maintaining its integrity is a prerequisite for survival. To ensure proper function, transmembrane signal-transducing systems, such as two-component systems (TCS) and extracytoplasmic function (ECF) factors, closely monitor its condition and respond to harmful perturbations. Both systems consist of a transmembrane sensor protein (histidine kinase or antifactor, respectively) and a corresponding cytoplasmic transcriptional regulator (response regulator or factor, respectively) that mediates the cellular response through differential gene expression. The regulatory network of the cell envelope stress response is well studied in the gram-positive model organism Bacillus subtilis. It consists of at least two ECF factors and four two-component systems. In this study, we describe the corresponding network in a close relative, Bacillus licheniformis. Based on sequence homology, domain architecture, and genomic context, we identified five TCS and eight ECF factors as potential candidate regulatory systems mediating cell envelope stress response in this organism. We characterized the corresponding regulatory network by comparative transcriptomics and regulon mining as an initial screening tool. Subsequent in-depth transcriptional profiling was applied to define the inducer specificity of each identified cell envelope stress sensor. A total of three TCS and seven ECF factors were shown to be induced by cell envelope stress in B. licheniformis. We noted a number of significant differences, indicative of a regulatory divergence between the two Bacillus species, in addition to the expected overlap in the respective responses.