Bacilysocin, a Novel Phospholipid Antibiotic Produced by Bacillus subtilis 168

Abstract

We have found a novel phospholipid antibiotic (named bacilysocin) which accumulates within (or associates with) the cells of Bacillus subtilis 168 and determined the structure by nuclear magnetic resonance and mass spectrometry analyses. The structure of bacilysocin elucidated was 1-(12-methyltetradecanoyl)-3-phosphoglyceroglycerol. Bacilysocin demonstrated antimicrobial activity, especially against certain fungi. Production of bacilysocin commenced immediately after growth ceased and before the formation of heat-resistant spores. The production of bacilysocin was completely blocked when the ytpA gene, which encodes a protein homologous to lysophospholipase, was disrupted, but blockage of the ytpA gene did not significantly affect growth. Sporulation was also impaired, with a 10-fold reduction in heat-resistant spore titers being detected. Since the ytpA disruptant actually lacked phospholipase activity, we propose that the YtpA protein functions as an enzyme for the biosynthesis of bacilysocin.The gram-positive bacterium Bacillus subtilis produces a large number of antibiotics, which are classified as ribosomal or nonribosomal. Nonribosomally synthesized circular oligopeptides that contain a fatty acid chain exhibit potent antibacterial or antifungal activity, as represented by surfactin, the iturinic group, and fengycin (16). B. subtilis 168 is the beststudied strain in the genus Bacillus, the genome of which was completely sequenced in 1997. Strain 168 is known to produce three ribosomal antibiotics, TasA (12), subtilosin (1), sublancin (10), and two nonribosomal antibiotics, surfactin (14) and bacilysin (9). The production of other antibiotics by strain 168 has also been predicted on the basis of genome sequence analysis, as exemplified by plipastatin (13). The ribosomal peptide antibiotics are synthesized during active growth, while nonribosomal ones are synthesized after growth has ceased. The role of antibiotic production for the producing organism is still under speculation. The best-accepted theory is that nonribosomal antibiotics may play a role in competition with other microorganisms during spore germination (for a review, see references 5 and 16). The detection of novel antibiotics produced by B. subtilis 168 would therefore be helpful in providing an understanding of the intrinsic (if any) role of antibiotics in the life cycle of this organism. In the present paper, we describe the isolation and identification of a new antibiotic (named bacilysocin) produced by B. subtilis 168. MATERIALS AND METHODSStrain, media, and culture conditions. B. subtilis strain 168 (trpC2) was precultured at 30°C for 24 h in NG medium (2) supplemented with 50 g of tryptophan per ml, and then 0.1 ml of the resulting culture was inoculated into 10 ml of fresh NG medium, followed by incubation under shaking at 30°C. The titers of the heat-resistant spores were determined by heating the cultures for 10 min at 80°C and then plating them onto an NG agar plate.Detection and bioassay of bacilysocin. Bacilysocin was extracted wit...

New Glycerides from a Two-Sponge Association ofJaspissp. andPoecillastrasp.

“…Furthermore, HMBC correlations from H-1a (δ 4.16) and H-1b (δ 4.09) to the carbonyl carbon at C-1 (δ 174.0) confirmed the ester linkage in the molecule. The presence of phosphate group was secured from the careful comparison of obtained NMR data with those reported (Tamehiro et al, 2002). The absolute configuration at C-2 was presumed to be R (vide infra).…”

“…Bacilysocin, a novel lysophosphatidylglycerol, isolated from Gram-positive bacteria (Bacillus subtilis), is reported to possess antimicrobial activity against bacteria and certain fungi (Tamehiro et al, 2002). One recent study showed that lysophosphatidylglycerols inhibit β-glucuronidase and can be used for prevention of colon and bladder cancers and also as drugs (oral or topical), health foods, and cosmetics for inhibiting unpleasant body odor (Shiojiri, 2007).…”

“…3) (De et al, 1993;Tamehiro et al, 2002). Glycerol 3-phosphate upon esterification with the first fatty acyl-CoA gives 1-acylglycerol 3-phosphate, which undergoes esterification with the second fatty acyl-CoA to form phosphatidic acid (1,2-diacylglycerol 3-phosphate).…”

“…This phosphatidic acid will be converted into phosphatidylphosphoglycerate on addition of a glycerol 3-phosphate molecule, which forms phosphatidylglycerol upon removal of a phosphate molecule. Enzyme lysophospholipase carries out hydrolysis of phosphatidylglycerol to yield lysophosphatidylglycerol (De et al, 1993;Tamehiro et al, 2002).…”

Characterization of a Novel Esterase YkoN from Bacillus subtilis Marburg

“…Among these the amino acid sequence of the product (YkoN) of ykoN has a weak homology with that of ugtP (ypfP) that is responsible for the synthesis of glucosylated diacylglycerates in this organism 5,7) , however, YkoN is found not to be involved in the synthesis of glucosylated diacylglycerates. The amino acid sequence of YkoN has a pentapeptide sequence (Gly-Gly-Ser-Met-Gly) that is compatible with the Gly/Ala-X-Ser-X-Gly motif of lipase/esterase as those shown among the microbial lipolytic enzymes, LipA, LipB, YtpA, and p-nitrobenzyl esterase (PnbA) of B. subtilis and lysophospholipase L2 (PldB) and Aes of E. coli [8][9][10][11][12][13][14] . We have purified His- Tagged YkoN and have shown that YkoN has an esterase activity.…”