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Structure-activity analysis of quorum-sensing signaling peptides from Streptococcus mutans.

ABSTRACT: Streptococcus mutans secretes and utilizes a 21-amino-acid signaling peptide pheromone to initiate quorum sensing for genetic competence, biofilm formation, stress responses, and bacteriocin production. In this study, we designed and synthesized a series of truncated peptides and peptides with amino acid substitutions to investigate their structure-activity relationships based on the three-dimensional structures of S. mutans wild-type signaling peptide UA159sp and C-terminally truncated peptide TPC3 from mutant JH1005 defective in genetic competence. By analyzing these peptides, we demonstrated that the signaling peptide of S. mutans has at least two functional domains. The C-terminal structural motif consisting of a sequence of polar hydrophobic charged residues is crucial for activation of the signal transduction pathway, while the core alpha-helical structure extending from residue 5 to the end of the peptide is required for receptor binding. Peptides in which three or more residues were deleted from the C terminus did not induce genetic competence but competitively inhibited quorum sensing activated by UA159sp. Disruption of the amphipathic alpha-helix by replacing the Phe-7, Phe-11, or Phe-15 residue with a hydrophilic residue resulted in a significant reduction in or complete loss of the activity of the peptide. In contrast to the C-terminally truncated peptides, these peptides with amino acid substitutions did not compete with UA159sp to activate quorum sensing, suggesting that disruption of the hydrophobic face of the alpha-helical structure results in a peptide that is not able to bind to the receptor. This study is the first study to recognize the importance of the signaling peptide C-terminal residues in streptococcal quorum sensing.

SUBMITTER: Syvitski RT

PROVIDER: S-EPMC1797343 | biostudies-literature | 2007 Feb

REPOSITORIES: biostudies-literature

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Structure-activity analysis of quorum-sensing signaling peptides from Streptococcus mutans.

Syvitski Raymond T RT Tian Xiao-Lin XL Sampara Kamal K Salman Alan A Lee Song F SF Jakeman David L DL Li Yung-Hua YH

Journal of bacteriology 20060825 4

Streptococcus mutans secretes and utilizes a 21-amino-acid signaling peptide pheromone to initiate quorum sensing for genetic competence, biofilm formation, stress responses, and bacteriocin production. In this study, we designed and synthesized a series of truncated peptides and peptides with amino acid substitutions to investigate their structure-activity relationships based on the three-dimensional structures of S. mutans wild-type signaling peptide UA159sp and C-terminally truncated peptide ...[more]

PMID: 16936029

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Project description:The genus Streptococcus encompasses a large bacterial taxon that commonly colonizes mucosal surfaces of vertebrates and is capable of disease etiologies originating from diverse body sites, including the respiratory, digestive, and reproductive tracts. Identifying new modes of treating infections is of increasing importance, as antibiotic resistance has escalated. Streptococcus mutans is an important opportunistic pathogen that is an agent of dental caries and is capable of systemic diseases such as endocarditis. As such, understanding how it regulates virulence and competes in the oral niche is a priority in developing strategies to defend from these pathogens. We determined that S. mutans UA159 possesses a bona fide short hydrophobic peptide (SHP)/Rgg quorum-sensing system that regulates a specialized biosynthetic operon featuring a radical-SAM (S-adenosyl-l-methionine) (RaS) enzyme and produces a ribosomally synthesized and posttranslationally modified peptide (RiPP). The pairing of SHP/Rgg regulatory systems with RaS biosynthetic operons is conserved across streptococci, and a locus similar to that in S. mutans is found in Streptococcus ferus, an oral streptococcus isolated from wild rats. We identified the RaS-RiPP product from this operon and solved its structure using a combination of analytical methods; we term these RiPPs tryglysin A and B for the unusual Trp-Gly-Lys linkage. We report that tryglysins specifically inhibit the growth of other streptococci, but not other Gram-positive bacteria such as Enterococcus faecalis or Lactococcus lactis We predict that tryglysin is produced by S. mutans in its oral niche, thus inhibiting the growth of competing species, including several medically relevant streptococci.IMPORTANCE Bacteria interact and compete with a large community of organisms in their natural environment. Streptococcus mutans is one such organism, and it is an important member of the oral microbiota. We found that S. mutans uses a quorum-sensing system to regulate production of a novel posttranslationally modified peptide capable of inhibiting growth of several streptococcal species. We find inhibitory properties of a similar peptide produced by S. ferus and predict that these peptides play a role in interspecies competition in the oral niche.

Project description:Streptococcus pneumoniae 5'-methylthioadenosine/S-adenosylhomocysteine hydrolase (MTAN) catalyzes the hydrolytic deadenylation of its substrates to form adenine and 5-methylthioribose or S-ribosylhomocysteine (SRH). MTAN is not found in mammals but is involved in bacterial quorum sensing. MTAN gene disruption affects the growth and pathogenicity of bacteria, making it a target for antibiotic design. Kinetic isotope effects and computational studies have established a dissociative S(N)1 transition state for Escherichia coli MTAN, and transition state analogues resembling the transition state are powerful inhibitors of the enzyme [Singh, V., Lee, J. L., Núñez, S., Howell, P. L., and Schramm, V. L. (2005) Biochemistry 44, 11647-11659]. The sequence of MTAN from S. pneumoniae is 40% identical to that of E. coli MTAN, but S. pneumoniae MTAN exhibits remarkably distinct kinetic and inhibitory properties. 5'-Methylthio-Immucillin-A (MT-ImmA) is a transition state analogue resembling an early S(N)1 transition state. It is a weak inhibitor of S. pneumoniae MTAN with a K(i) of 1.0 microM. The X-ray structure of S. pneumoniae MTAN with MT-ImmA indicates a dimer with the methylthio group in a flexible hydrophobic pocket. Replacing the methyl group with phenyl (PhT-ImmA), tolyl (p-TolT-ImmA), or ethyl (EtT-ImmA) groups increases the affinity to give K(i) values of 335, 60, and 40 nM, respectively. DADMe-Immucillins are geometric and electrostatic mimics of a fully dissociated transition state and bind more tightly than Immucillins. MT-DADMe-Immucillin-A inhibits with a K(i) value of 24 nM, and replacing the 5'-methyl group with p-Cl-phenyl (p-Cl-PhT-DADMe-ImmA) gave a K(i) value of 0.36 nM. The inhibitory potential of DADMe-Immucillins relative to the Immucillins supports a fully dissociated transition state structure for S. pneumoniae MTAN. Comparison of active site contacts in the X-ray crystal structures of E. coli and S. pneumoniae MTAN with MT-ImmA would predict equal binding, yet most analogues bind 10(3)-10(4)-fold more tightly to the E. coli enzyme. Catalytic site efficiency is primarily responsible for this difference since k(cat)/K(m) for S. pneumoniae MTAN is decreased 845-fold relative to that of E. coli MTAN.

Dataset Information

Structure-activity analysis of quorum-sensing signaling peptides from Streptococcus mutans.

Publications

Structure-activity analysis of quorum-sensing signaling peptides from Streptococcus mutans.

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