Biosynthesis of teichuronic acid in the bacterial cell wall. Purification and characterization of the glucosyltransferase of Micrococcus luteus.
This report describes what is, to our knowledge, the first purification to near homogeneity of an enzyme involved in the biosynthesis of the teichuronic acid of Micrococcus luteus cell walls. The glucosyltransferase of M. luteus, which participates in the biosynthesis of teichuronic acid, was solubilized from cytoplasmic membrane fragments by extraction with buffer solutions containing the detergents Thesit (dodecyl alcohol polyoxyethylene ether; 1 mg/ml) and 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (0.5 mg/ml). The detergent-solubilized enzyme was purified 150-fold, with a recovery of 13% by adsorbent column chromatography, ion-exchange chromatography, gel filtration, and preparative nondenaturing gradient polyacrylamide gel electrophoresis. On the basis of its mobility on native gradient gel, the glucosyltransferase was estimated to have a molecular mass of 440 kDa. The purified native enzyme was a multisubunit protein consisting of subunits of two sizes; their molecular masses were determined to be 52.5 and 54 kDa, respectively, by observation of the mobility of the protein bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric point of the enzyme was approximately 5.
Deng
L
L
Department of Biochemistry, College of Biological Sciences, University of Minnesota, St. Paul 55108, USA.
Anderson
J S
JS
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000302:isolation & purification; D002473:Cell Wall; D003902:Detergents; D005964:Glucosyltransferases / Q000302:isolation & purification*; D007526:Isoelectric Point; D008565:Membrane Proteins / Q000302:isolation & purification; D016982:Micrococcus luteus / Q000201:enzymology*; D008970:Molecular Weight; D014574:Uronic Acids / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D003902:Detergents; D008565:Membrane Proteins; D014574:Uronic Acids; C010788:teichuronic acid; D005964:Glucosyltransferases