Mutational Analysis of the sbo-alb Locus of Bacillus subtilis : Identification of Genes Required for Subtilosin Production and Immunity

Abstract

The Bacillus subtilis 168 derivative JH642 produces a bacteriocin, subtilosin, which possesses activity against Listeria monocytogenes. Inspection of the amino acid sequence of the presubtilosin polypeptide encoded by the gene sboA and sequence data from analysis of mature subtilosin indicate that the precursor subtilosin peptide undergoes several unique and unusual chemical modifications during its maturation process. The genes of the sbo-alb operon are believed to function in the synthesis and maturation of subtilosin. Nonpolar mutations introduced into each of the alb genes resulted in loss or reduction of subtilosin production. sboA, albA, and albF mutants showed no antilisterial activity, indicating that the products of these genes are critical for the production of active subtilosin. Mutations in albB, -C, and -D resulted in reduction of antilisterial activity and decreased immunity to subtilosin, particularly under anaerobic conditions. A new gene, sboX, encoding another bacteriocin-like product was discovered residing in a sequence overlapping the coding region of sboA. Construction of an sboX-lacZ translational fusion and analysis of its expression indicate that sboX is induced in stationary phase of anaerobic cultures of JH642. An in-frame deletion of the sboX coding sequence did not affect the antilisterial activity or production of or immunity to subtilosin. The results of this investigation show that the sbo-alb genes are required for the mechanisms of subtilosin synthesis and immunity.

Thuricin CD, a posttranslationally modified bacteriocin with a narrow spectrum of activity against Clostridium difficile

“…TrnC exhibited sequence similarity to conserved domains of the Fe-S oxidoreductase arylsulfatase regulatory proteins (AslB), and TrnD had sequences similar to the MoaA family of molybdenum cofactor enzymes. Radical SAM enzymes rarely occur in bacteriocin gene clusters, but they are included in those of subtilosin A and propiocin F (26,27). Sequence alignments indicated that AlbA of subtilosin A exhibits only 19% and 17% identity to TrnC and TrnD, respectively, although AlbA exhibits some features of each individual thuricin CD protein.…”

Bacteria Exhibiting Antimicrobial Activities; Screening for Antibiotics and the Associated Genetic Studies

“…PCR sequences were identified using the basic local alignment search tool and GenBank nucleotide data bank from the National Center for Biotechnology Information, Bethesda, MD, USA (http://www.ncbi.nlm.nih.gov). sboX, encoded a bacteriocinlike product, a new gene with an unknown function, in strain 168 [19], which resides in an open reading frame overlapping the coding region of sbo. While the expression of sboX would result in a 22-amino-acid curtailed peptide in W23-like strains compared to the peptide produced by 168-like strains, which makes it unlikely that SboX is produced by W23-like strains.…”

“…The spectrum of activity was investigated and proved antagonism against a wide range of bacteria including Gram-positive and Gram-negative bacteria and both aerobes and anaerobes [18]. The production of mature subtilosin is based on the expression of the sbo-alb *Address correspondence to this author at the Department of Microbiology and Molecular Genetics, University of the Punjab, Quaid-e-Azam Campus, Lahore, Pakistan; Tel: +92-42-99238530; Fax +92-42-99230481: E-mail: shahida@mmg.pu.edu.pk, genetic@brain.net.pk gene cluster encompassing the subtilosin structural gene sbo and genes involved in posttranslational modification, processing of presubtilosin and immunity [17,19].…”

Isolation of a Variant of Subtilosin A with Hemolytic Activity

“…AlbB, AlbC, and AlbD are required for subtilosin A immunity. AlbB was postulated to be an immunity peptide and AlbC to be involved in subtilosin A export (37). The albABCDEF genes are transcribed from the sboA promoter despite a putative factor-independent transcription terminator present between sboA and albA.…”

“…The sboA gene, encoding the 43-amino-acid precursor of subtilosin A, was identified with the downstream albABCDEF genes, which are required for processing and modification of the precursor as well as immunity to subtilosin A (37,38). AlbA and AlbF are essential for subtilosin A production and are likely involved in modification and processing of the precursor, respectively.…”