Biosynthesis of the lantibiotic mersacidin: organization of a type B lantibiotic gene cluster

Abstract

The biosynthetic gene cluster (12.3 kb) of mersacidin, a lanthionine-containing antimicrobial peptide, is located on the chromosome of the producer, Bacillus sp. strain HIL Y-85,54728 in a region that corresponds to 348 degrees on the chromosome of Bacillus subtilis 168. It consists of 10 open reading frames and contains, in addition to the previously described mersacidin structural gene mrsA (G. Bierbaum, H. Brötz, K.-P. Koller, and H.-G. Sahl, FEMS Microbiol. Lett. 127:121-126, 1995), two genes, mrsM and mrsD, coding for enzymes involved in posttranslational modification of the prepeptide; one gene, mrsT, coding for a transporter with an associated protease domain; and three genes, mrsF, mrsG, and mrsE, encoding a group B ABC transporter that could be involved in producer self-protection. Additionally, three regulatory genes are part of the gene cluster, i.e., mrsR2 and mrsK2, which encode a two-component regulatory system which seems to be necessary for the transcription of the mrsFGE operon, and mrsR1, which encodes a protein with similarity to response regulators. Transcription of mrsA sets in at early stationary phase (between 8 and 16 h of culture).

FIG. 1

Mersacidin biosynthetic gene cluster. (A) Inserts of sequencing clones. (B) Organization of the gene cluster. The arrows indicate relative directions of transcription. (C) Inserts of plasmids constructed for transcription experiments with mrsFGE.

FIG. 2

Comparison of the sequences of MrsD, MutD, and EpiD from the mutacin III and epidermin biosynthesis gene clusters (1, 38). The asterisks indicate identical residues, and residues with high and low degrees of conservation are indicated by colons and dots, respectively.

FIG. 3

Conserved motifs of the N termini of the LanM proteins. The asterisks indicate identical residues, and residues with high and low degrees of conservation are indicated by colons and dots, respectively.

FIG. 4

Northern blot analysis of the time course of mrsA transcription. Lanes: 1, size standard (the values to the left are sizes in kilobases); 2, 8 h of cultivation; 3, 16 h of cultivation; 4, 24 h of cultivation; 5, 32 h of cultivation.

FIG. 5

Time course of production of antibacterial activity (diameter of inhibition zone against M. luteus, in centimeters (Δ) and growth (□) of the wild-type producer strain (A) and Bacillus sp. strain HIL Y-85,54728 Rec1 (mrsA::Erm) (B).

FIG. 6

Northern blot analysis of the transcription of mrsFGE employing an mrsF probe. Lanes: 1, size standard (the values to the left are sizes in kilobases); 2, wild-type producer Bacillus sp. strain HIL Y-85,54728; 3, B. subtilis W23 harboring pCC2; 4, B. subtilis W23 harboring pKRFGE2.

FIG. 7

Alignment of the C-terminal part of the leader sequences of lantibiotic prepeptides harboring a GG-type cleavage site with an internal fragment of the mersacidin prepeptide. Amino acid residues that are not conserved are in lowercase, whereas residues that are conserved in five or more leader peptides are in uppercase and boxed (11, 30, 33, 40, 42, 50).