Expression of a Streptococcus mutans glucosyltransferase gene in Escherichia coli

J. P. Robeson, Raul G Barletta, R. Curtiss

Research output: Contribution to journalArticle

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Abstract

Chromosomal DNA from S. mutans strain UAB90 (serotype c) was cloned into E. coli K-12. The clone bank was screened for any sucrose-hydrolyzing activity by selection for growth on raffinose in the presence of isopropyl-β-D-thiogalactoside. A clone expressing on S. mutans glycosyltransferase was identified. The S. mutans DNA encoding this enzyme is a 1.73-kilobase fragment cloned into the HindIII site of plasmid pBR322. The authors designated the gene gtfA. The plamid-encoded gtfA enzyme, a 55,000-molecular-weight protein, is synthesized at 40% the level of pBR322-encoded β-lactamase in E. coli minicells. Using sucrose as substrate, the gtfA enzyme catalyzes the formation of fructose and a glucan with an apparent molecular weight of 1,500. They detected the gtfA protein in S. mutans cells with antibody raised against the cloned gtfA enzyme. Immunologically identical gtfA protein appears to be present in S. mutans cells of serotypes C, e, and f, and a cross-reacting protein was made by serotype b cells. Proteins from serotype a, g, and d S. mutans cells did not react with antibody to gtfA enzyme. The gtfA activity was present in the periplasmic space of E. coli clones, since 15% of the total gtfA activity was released by cold osmotic shock and the clones were able to grow on sucrose as sole carbon source.

Original languageEnglish (US)
Pages (from-to)211-221
Number of pages11
JournalJournal of bacteriology
Volume153
Issue number1
StatePublished - Jan 1 1983

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Glucosyltransferases
Streptococcus mutans
Escherichia coli
Clone Cells
Enzymes
Sucrose
Genes
Proteins
Molecular Weight
Raffinose
Isopropyl Thiogalactoside
Periplasm
Glycosyltransferases
Glucans
Antibodies
Protein S
DNA
Osmotic Pressure
Fructose
Plasmids

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

Cite this

Expression of a Streptococcus mutans glucosyltransferase gene in Escherichia coli. / Robeson, J. P.; Barletta, Raul G; Curtiss, R.

In: Journal of bacteriology, Vol. 153, No. 1, 01.01.1983, p. 211-221.

Research output: Contribution to journalArticle

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