Purification and properties of dextransucrase from Streptococcus mutans.
The dextransucrase (EC 2.4.1.5) activity from cell-free culture supernatants of Streptococcus mutans strain 6715 has been purified approximately 1,500-fold by ammonium sulfate precipitation, hydroxylapatite chromatography, and isoelectric focusing. The enzyme was eluted as a single peak of activity from hydroxylapatite, and isoelectric focusing of the resulting preparation gave a single band of dextransucrase activity which focused at a pH of 4.0. The final enzyme preparation contained two distinct, enzymatically active proteins as judged by assay in situ after polyacrylamide gel electrophoresis. One of the proteins represented 90% of the total dextransucrase activity and 53% of the total protein. The molecular weight of the enzyme was estimated by gel filtration to be 94,000. The temperature optimum of the enzyme was broad (34 to 42 C) and its pH range was rather narrow, with optimal activity at pH 5.5. The K(m) for sucrose was 3 mM, and fructose competitively inhibited the enzyme reaction with a K(i) of 27 mM.
D000645:Ammonium Sulfate; D001345:Autoradiography; D001426:Bacterial Proteins / Q000032:analysis; D002250:Carbon Radioisotopes; D002474:Cell-Free System; D011232:Chemical Precipitation; D002845:Chromatography; D002850:Chromatography, Gel; D003911:Dextrans / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D005632:Fructose / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism; D005964:Glucosyltransferases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006882:Hydroxyapatites; D007525:Isoelectric Focusing; D008970:Molecular Weight; D013291:Streptococcus / Q000201:enzymology*; D013395:Sucrose / Q000378:metabolism; D013696:Temperature
D001426:Bacterial Proteins; D002250:Carbon Radioisotopes; D003911:Dextrans; D006882:Hydroxyapatites; D005632:Fructose; D013395:Sucrose; D005964:Glucosyltransferases; D005947:Glucose; D000645:Ammonium Sulfate