Signal Peptide-Dependent Protein Transport in Bacillus subtilis : a Genome-Based Survey of the Secretome

Abstract

SUMMARY One of the most salient features of Bacillus subtilis and related bacilli is their natural capacity to secrete a variety of proteins into their environment, frequently to high concentrations. This has led to the commercial exploitation of bacilli as major “cell factories” for secreted enzymes. The recent sequencing of the genome of B. subtilis has provided major new impulse for analysis of the molecular mechanisms underlying protein secretion by this organism. Most importantly, the genome sequence has allowed predictions about the composition of the secretome, which includes both the pathways for protein transport and the secreted proteins. The present survey of the secretome describes four distinct pathways for protein export from the cytoplasm and approximately 300 proteins with the potential to be exported. By far the largest number of exported proteins are predicted to follow the major “Sec” pathway for protein secretion. In contrast, the twin-arginine translocation “Tat” pathway, a type IV prepilin-like export pathway for competence development, and ATP-binding cassette transporters can be regarded as “special-purpose” pathways, through which only a few proteins are transported. The properties of distinct classes of amino-terminal signal peptides, directing proteins into the various protein transport pathways, as well as the major components of each pathway are discussed. The predictions and comparisons in this review pinpoint important differences as well as similarities between protein transport systems in B. subtilis and other well-studied organisms, such as Escherichia coli and the yeast Saccharomyces cerevisiae. Thus, they may serve as a lead for future research and applications.

Characterization of new l,d-endopeptidase gene product CwlK (previous YcdD) that hydrolyzes peptidoglycan in Bacillus subtilis

“…Since from the SignalP algorithm it is predicted that CwlK seems to have a secretory signal peptide, it is possible that CwlK is secreted and localized outside of the cell. On the other hand, because it is also predicted that this protein seems to have a lipoprotein signal peptide described by Tjalsma et al (2000), CwlK may be localized in membrane as a lipoprotein. In order to clarify the role of CwlK, we determined the localization of CwlK with YCDDd (cwlK::erm) and the wild-type strain during the vegetative growth phase by SDS-PAGE and zymography.…”

“…The cwlK gene encodes a polypeptide consisting of 167 amino acid residues and its N-terminal region contains a signal sequence based on the SignalP (http://www.cbs.dtu.dk/services/SignalP/) algorithm, and the signal sequence is predicted to be MNLPAKTFVILCILFLLDLCFSYIRH 26 ↓E 27 (the arrow indicates the cleavage point and the numbers are with respect to the N-terminal amino acid residue). Tjalsma et al (2000) also described that the signal sequence of CwlK is assumed to be The wild-type B. subtilis is rod-shaped, and exists as short chains in the vegetative growth phase and as short rods in the stationary phase. Several previous reports indicated that autolysins in B. subtilis can induce lysis, and play roles in cell separation, cell elongation, motility, cell wall turnover, etc.…”

The neomuran origin of archaebacteria, the negibacterial root of the universal tree and bacterial megaclassification.

“…One particular phylum alone, the Proteobacteria, made the further addition of the SecB chaperone to reduce the problem of denaturation and degradation of proteins prior to secretion. I suggest that this problem became particularly acute in actinobacteria, which are often thermophiles (never hyperthermophiles) that secrete an unusually large number of proteins or peptides ; pronounced protein secretion is a basic characteristic of posibacteria -Bacillus subtilis secretes over 300 (Tjalsma et al, 2000) ; its more-complex SRP has helices 1-4 like neomura. These two features of their lifestyle may explain why the ancestral actinobacterium evolved proteasomes for the degradation of misfolded or denatured proteins.…”

“…not all, secretory proteins follow this post-translational pathway. In Bacillus subtilis (and, very likely, other posibacteria), however, only a minority of secretory enzymes use the SecA post-translational mechanism ; the great majority are probably secreted co-translationally by the SRP mechanism (Tjalsma et al, 2000). Neomura, however, do not have SecA and secrete essentially all proteins with a cleavable signal sequence co-translationally.…”

Heterologous production and secretion of Clostridium perfringens β-toxoid in closely related Gram-positive hosts

“…The C. perfringens ␤-toxoid native signal sequence is a consensus Sec-type signal sequence, and in theory compatible with the B. subtilis secretion system (Tjalsma et al, 2000). Swapping the ␤-toxoid signal sequence with the signal sequence of a well secreted Bacillus protein (AmyQ) did not increase its presence in the growth medium.…”