The specific genes for lantibiotic mutacin II biosynthesis in Streptococcus mutans T8 are clustered and can be transferred en bloc - PubMed (original) (raw)

The specific genes for lantibiotic mutacin II biosynthesis in Streptococcus mutans T8 are clustered and can be transferred en bloc

P Chen et al. Appl Environ Microbiol. 1999 Mar.

Abstract

Mutacin II is a ribosomally synthesized peptide lantibiotic produced by group II Streptococcus mutans. DNA sequencing has revealed that the mutacin II biosynthetic gene cluster consists of seven specific open reading frames: a regulator (mutR), the prepromutacin structural gene (mutA), a modifying protein (mutM), an ABC transporter (mutT), and an immunity cluster (mutFEG). Transformations of a non-mutacin-producing strain, S. mutans UA159, and a mutacin I-producing strain, S. mutans UA140, with chromosomal DNA from S. mutans T8 with an aphIII marker inserted upstream of the mutacin II structural gene yielded transformants producing mutacin II and mutacins I and II, respectively.

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Figures

FIG. 1

Deferred-antagonism assay of mutacin phenotypes in selected S. mutans strains. 1, T8 wild type; 2 to 4, UA159 transformants; 5, UA159 wild type; 6, indicator strain S. sobrinus OMZ176. Mutacin production was indicated by the inhibition zones around the producing strains (strain T8 and UA159 transformants) in the lawn of S. sobrinus OMZ176. The absence of such an inhibition zone indicated a lack of mutacin production (parental strain UA159 and the negative control, S. sobrinus OMZ176).

FIG. 2

Restriction and Southern blot analysis of S. mutans mutacin-producing and non-mutacin-producing strains and corresponding transformants. Chromosomal DNA was digested with _Hin_dIII, and the resultant fragments were resolved on a 0.65% agarose gel in Tris-borate-EDTA buffer and photographed under UV light (A). The gel was transferred to a nylon membrane and probed with part of the mutacin II biosynthetic operon (mutAM) (B). Lanes: 1, T8 wild type; 2, UA159 wild type; 3 to 5, UA159 transformants; 6, UA140 wild type, 7 to 9, UA140 transformants. DNA marker sizes are indicated on the left side of each panel, in kilobases.

FIG. 3

Electrospray ionization mass spectroscopic analysis of purified mutacin isolated from S. mutans UA159 transformants by a previously described method (15). The molecular mass indicated by multiple charged ions was calculated as 3,245.4 ± 0.6 Da, which is identical to that of mutacin II from S. mutans T8 (15).

FIG. 4

Deferred-antagonism assay of the mutacin I and II phenotypes in selected S. mutans strains. (A) When S. mutans T8 was used as the indicator strain, the inhibition zones around S. mutans UA140 wild type (5) and its transformants (2 to 4) indicated mutacin I production in these strains, while the absence of such an inhibition zone around T8 wild type (1) indicated that mutacin I was not produced by this strain. (B) Mutacin II production by T8 wild type (1) and UA140 transformants (2 to 4) was indicated by their ability to inhibit growth of S. aureus MSSA4.

FIG. 5

Genetic organization of the mutacin II biosynthetic gene cluster. The cluster consists of seven specific genes in the following order: regulator gene (mutR), structural gene (mutA), modifying enzyme gene (mutM), transporter and leader peptidase gene (mutT), and accessory self-protection genes (mutFEG). Upstream of mutR is a silent transposase gene (tra).

References

1. Augustin J, Rosenstein R, Wieland B, Schneider U, Norbert S, Engelke G, Entain K D, Gotz F. Genetic analysis of epidermin biosynthetic genes and epidermin-negative mutants of Staphylococcus epidermidis. Eur J Biochem. 1992;204:1149–1154. -PubMed
1. Braun V, Hundsberger T, Leukel P, Sauerborn M, von Eichel-Streiber C. Definition of the single integration site of the pathogenicity locus in Clostridium difficile. Gene. 1996;181:29–38. -PubMed
1. Buchman G W, Banerje S, Hansen J N. Structure, expression, and evolution of a gene encoding the precursor of nisin, a small protein antibiotic. J Biol Chem. 1988;263:16260–16266. -PubMed
1. Caufield P W, Childers N K, Allen D N, Hansen J B. Distinct bacteriocin groups correlate with different groups of Streptococcus mutans plasmids. Infect Immun. 1985;48:51–56. -PMC -PubMed
1. Chen P, Novak J, Kirk M, Barnes S, Qi F, Caufield P W. Structure-activity study of the lantibiotic mutacin II from Streptococcus mutans T8 by a gene replacement strategy. Appl Environ Microbiol. 1998;64:2335–2340. -PMC -PubMed

The specific genes for lantibiotic mutacin II biosynthesis in Streptococcus mutans T8 are clustered and can be transferred en bloc - PubMed (original) (raw)