The endogenous Bacillus subtilis (natto) plasmids pTA1015 and pTA1040 contain signal peptidase‐encoding genes: identification of a new structural module on cryptic plasmids

Abstract

Various strains of Bacillus subtilis (natto) contain small cryptic plasmids that replicate via the rolling-circle mechanism. Like plasmids from other Gram-positive bacteria, these plasmids are composed of several distinct structural modules. A new structural module was identified on the B. subtilis plasmids pTA1015 and pTA1040. It is composed of two genes: one specifies an unidentified protein with a putative signal peptide; and the other (sipP) specifies a functional type 1 signal peptidase (SPase). The homologous, but non-identical, sipP genes of the two plasmids are the first identified plasmid-specific SPase-encoding genes. With respect to structure and activity, the corresponding enzymes (denoted SipP) are highly similar to the chromosomally encoded SPase, SipP, of B. subtillis and several newly identified SPases of other bacilli. Our findings suggest that plasmid-encoded SPases have evolved because, of under certain conditions, SPase can be a limiting factor for protein secretion in B. subtilis.

The Potential Active Site of the Lipoprotein-specific (Type II) Signal Peptidase of Bacillus subtilis

“…Amplified fragments were subsequently cleaved with SalI and EcoRI and ligated into the corresponding sites of pGDL48 (26). Consequently, the wild-type or mutant lsp genes on pGDL150 -152 were transcribed from the constitutive promoter of the Em r gene present on pGDL48.…”

The chemistry and enzymology of the type I signal peptidases

“…1A). The latter feature is shared by the signal peptidase of the cyanobacterium Phonnidium laminosum (22 kDa; Packer et al, 1995), and all known signal peptidases from Gram positive bacteria, which include enzymes from Bacillus subtilis (van Dijl et al, 1992), Bacillus amyloliquefaciens, Bacillus culdolyticus, and Bacillus lichenifonnis (for a recent compilation of sequences see Meijer et al, 1995), Staphylococcus aureus (SpsB; Cregg et al, 1996), and Mycobacterium tuberculosis (Philipp et al, 1996). All Bacillus signal peptidases and the S. uureus signal peptidase consist of 21 kDa polypeptides whereas the M. tuberculosis gene encodes a 32 kDa polypeptide.…”

“…Interestingly, both in B. subtilis and B. amyloliquefaciens, chromosomal genes were identified for two homologous, but nonidentical, type I signal peptidases, denoted Sips and SipT (van Dijl et al, 1992;Meijer et al, 1995;Hoang & Hofemeister 1995;GenBank accession #U45883). In addition, a third chromosomal gene for a homologous type I signal peptidase (SipU) was identified in B. subtilis (Akagawa et al, 1995; H. Tjalsma, S. Bron & J. M. van Dijl, unpubl.…”

Detergent-independent in vitro activity of a truncated Bacillus signal peptidase

“…Type I signal peptidases (SPases) remove these signal peptides during, or shortly after, pre-protein translocation across the cytoplasmic membrane, thereby releasing the mature proteins from the trans side of the membrane (for reviews, see Pugsley, 1993 ;Dalbey et al, 1997). The largest number of type I SPases has, thus far, been found in the Gram-positive eubacterium Bacillus subtilis, which contains five paralogous, chromosomally encoded enzymes of this type (SipS, SipT, SipU, SipV and SipW) (van Dijl et al, 1992 ;Bolhuis et al, 1996 ;Tjalsma et al, 1997Tjalsma et al, , 1998 plasmid-encoded type I SPases, denoted SipP (Meijer et al, 1995 ;Tjalsma et al, 1999a). Even though the presence of paralogous SPases is not unusual in eubacteria, archaea and eukaryotes, the number of paralogous SPases in B. subtilis is unusually high, suggesting that some of these SPases may have specialized functions.…”