gramicidin-a and surfactin-peptide

In Bacillus species, starvation leads to the activation of a number of processes that affect the ability to survive during periods of nutritional stress. Activities that are induced include the development of genetic competence, sporulation, the synthesis of degradative enzymes, motility, and antibiotic production. The genes that function in these processes are activated during the transition from exponential to stationary phase and are controlled by mechanisms that operate primarily at the level of transcription initiation. One class of genes functions in the synthesis of special metabolites such as the peptide antibiotics tyrocidine and gramicidin S as well as the cyclic lipopeptide surfactin. These genes include the grs and tyc operons in Bacillus brevis, which encode gramicidin S synthetase and tyrocidine synthetase, respectively, and the srfA operon of Bacillus subtilis which encodes the enzymes of the surfactin synthetase complex. Peptide antibiotic biosynthesis genes are regulated by factors as diverse as the early sporulation gene product Spo0A, the transition-state regulator AbrB, and gene products (ComA, ComP, and ComQ) required for the initiation of the competence developmental pathway.

Topics: Amino Acid Isomerases; Bacillus; Bacillus subtilis; Bacterial Proteins; Base Sequence; DNA-Binding Proteins; Enzyme Induction; Gene Expression Regulation, Bacterial; Gramicidin; Lipopeptides; Membrane Proteins; Models, Biological; Molecular Sequence Data; Multienzyme Complexes; Operon; Peptide Synthases; Peptides, Cyclic; Spores, Bacterial; Transferases; Tyrocidine

Several species of the genus Bacillus produce peptide antibiotics which are synthesized either through a ribosomal or non-ribosomal mechanism. The antibiotics gramicidin, tyrocidine, and bacitracin are synthesized nonribosomally by the multienzyme thiotemplate mechanism. Surfactin and mycobacillin are also synthesized nonribosomally but by a mechanism that, apparently, is distinct from that of the multienzyme thiotemplate. Other antibiotics such as subtilin are gene encoded and are ribosomally synthesized. Molecular genetic and DNA sequence analysis have shown that biosynthesis genes for some antibiotics are clustered into polycistronic transcription units and are under the control of global regulatory systems that govern the expression of genes that are induced when Bacillus cells enter stationary phase of growth. Future experiments involving the molecular dissection of peptide antibiotic biosynthesis genes in Bacillus will be attempted in hopes of further examining the mechanism and regulation of antibiotic production.

Topics: Amino Acid Sequence; Anti-Bacterial Agents; Bacillus; Bacterial Proteins; Base Sequence; Gramicidin; Lipopeptides; Molecular Sequence Data; Mutation; Peptides, Cyclic; Ribosomes; Tyrocidine

Microbial colonization can be detrimental to the integrity of metal surfaces and lead to microbiologically influenced corrosion (MIC). Biocorrosion is a serious problem for aquatic and marine industries in the world. In Minnesota (USA), where this study was conducted, biocorrosion severely affects the maritime transportation industry. The anticorrosion activity of a variety of compounds, including chemical (magnesium peroxide) and biological (surfactin, capsaicin, and gramicidin) molecules were investigated as coating additives. We also evaluated a previously engineered, extremely stable, non-biocidal enzyme known to interfere in bacterial signaling, SsoPox (a quorum quenching lactonase). Experimental steel coupons were submerged in water from the Duluth Superior Harbor (DSH) for 8 weeks in the laboratory. Biocorrosion was evaluated by counting the number and the coverage of corrosion tubercles on coupons and also by ESEM imaging of the coupon surface. Three experimental coating additives significantly reduced the formation of corrosion tubercles: surfactin, magnesium peroxide and the quorum quenching lactonase by 31%, 36% and 50%, respectively. DNA sequence analysis of the V4 region of the bacterial 16S rRNA gene revealed that these decreases in corrosion were associated with significant changes in the composition of bacterial communities on the steel surfaces. These results demonstrate the potential of highly stable quorum quenching lactonases to provide a reliable, cost-effective method to treat steel structures and prevent biocorrosion.

Topics: Anti-Bacterial Agents; Bacteria; Biofilms; Capsaicin; Carboxylic Ester Hydrolases; Corrosion; Gramicidin; Lipopeptides; Magnesium Compounds; Minnesota; Peptides, Cyclic; Peroxides; Quorum Sensing; RNA, Ribosomal, 16S; Steel; Surface Properties; Water Microbiology

It was previously observed that the lipopeptide surfactants in surfactin (Srf) have an antagonistic action towards the highly potent antimicrobial cyclodecapeptide, gramicidin S (GS). This study reports on some of the molecular aspects of the antagonism as investigated through complementary electrospray ionization mass spectrometry techniques. We were able to detect stable 1:1 and 2:1 hetero-oligomers in a mixture of surfactin and gramicidin S. The noncovalent interaction between GS and Srf, with the proposed equilibrium: GS~Srf↔GS+Srf correlated to apparent K

Topics: Anti-Infective Agents; Binding Sites; Dimerization; Gramicidin; Hydrophobic and Hydrophilic Interactions; Lipopeptides; Peptides, Cyclic; Spectrometry, Mass, Electrospray Ionization

Antibiotic production as a defence mechanism is a characteristic of a wide variety of organisms. In natural evolutionary adaptation, cellular events such as sporulation, biofilm formation and resistance to antibiotics enable some micro-organisms to survive environmental and antibiotic stress conditions. The two antimicrobial cyclic peptides in this study, gramicidin S (GS) from Aneurinibacillus migulanus and the lipopeptide surfactin (Srf) from Bacillus subtilis, have been shown to affect both membrane and intercellular components of target organisms. Many functions, other than that of antimicrobial activity, have been assigned to Srf. We present evidence that an additional function may exist for Srf, namely that of a detoxifying agent that protects its producer from the lytic activity of GS. We observed that Srf producers were more resistant to GS and could be co-cultured with the GS producer. Furthermore, exogenous Srf antagonized the activity of GS against both Srf-producing and non-producing bacterial strains. A molecular interaction between the anionic Srf and the cationic GS was observed with circular dichroism and electrospray MS. Our results indicate that the formation of an inactive complex between GS and Srf supports resistance towards GS, with the anionic Srf forming a chemical barrier to protect its producer. This direct detoxification combined with the induction of protective stress responses in B. subtilis by Srf confers resistance toward GS from A. migulanus and allows survival in mixed cultures.

Topics: Anti-Bacterial Agents; Antibiosis; Bacillales; Circular Dichroism; Drug Resistance, Bacterial; Gramicidin; Inactivation, Metabolic; Lipopeptides; Peptides, Cyclic; Protein Binding; Spectrometry, Mass, Electrospray Ionization

The C-terminal thioesterase (TE) domains from nonribosomal peptide synthetases (NRPSs) catalyze the final step in the biosynthesis of diverse biologically active molecules. In many systems, the thioesterase domain is involved in macrocyclization of a linear precursor presented as an acyl-S-enzyme intermediate. The excised thioesterase domain from the tyrocidine NRPS has been shown to catalyze the cyclization of a peptide thioester substrate which mimics its natural acyl-S-enzyme substrate. In this work we explore the generality of cyclization catalyzed by isolated TE domains. Using synthetic peptide thioester substrates from 6 to 14 residues in length, we show that the excised TE domain from the tyrocidine NRPS can be used to generate an array of sizes of cyclic peptides with comparable kinetic efficiency. We also studied the excised TE domains from the NRPSs which biosynthesize the symmetric cyclic decapeptide gramicidin S and the cyclic lipoheptapeptide surfactin A. Both TE domains exhibit expected cyclization activity: the TE domain from the gramicidin S NRPS catalyzes head-to-tail cyclization of a decapeptide thioester to form gramicidin S, and the TE domain from the surfactin NRPS catalyzes stereospecific cyclization to form a macrolactone analogue of surfactin. With an eye toward generating libraries of cyclic molecules by TE catalysis, we report the solid-phase synthesis and TE-mediated cyclization of a small pool of linear peptide thioesters. These studies provide evidence for the general utility of TE catalysis as a means to synthesize a wide range of macrocyclic compounds.

Topics: Amino Acid Isomerases; Bacterial Proteins; Catalysis; Gramicidin; Lipopeptides; Lipoproteins; Multienzyme Complexes; Peptide Fragments; Peptide Synthases; Peptides, Cyclic; Protein Structure, Tertiary; Substrate Specificity; Thiolester Hydrolases

The deduced amino acid sequence of the gsp gene, located upstream of the 5' end of the gramicidin S operon (grs operon) in Bacillus brevis, showed a high degree of similarity to the sfp gene product, which is located downstream of the srfA operon in B. subtilis. The gsp gene complemented in trans a defect in the sfp gene (sfpO) and promoted production of the lipopeptide antibiotic surfactin. The functional homology of Gsp and Sfp and the sequence similarity of these two proteins to EntD suggest that the three proteins represent a new class of proteins involved in peptide secretion, in support of a hypothesis published previously (T. H. Grossman, M. Tuckman, S. Ellestad, and M. S. Osburne, J. Bacteriol. 175:6203-6211, 1993).

Topics: Amino Acid Sequence; Bacillus; Bacillus subtilis; Bacterial Proteins; Base Sequence; Gene Expression; Genes, Bacterial; Genetic Complementation Test; Gramicidin; Lipopeptides; Molecular Sequence Data; Operon; Peptides, Cyclic; Sequence Homology; Transferases (Other Substituted Phosphate Groups)