|
KMID : 0545120040140051075
|
|
Journal of Microbiology and Biotechnology
2004 Volume.14 No. 5 p.1075 ~ p.1080
|
|
|
Identification of Amino-Acids Residues for Key Role in Dextransucrase Activity of Leuconostoc mesenteroides B-742CB
|
|
Kimura A
|
|
|
Abstract
|
|
|
|
Dextransucrase (DSRB742) from Leuconostoc mesenteroides NRRL B-742CB is a glucosyltransferase that catalyzes the synthesis of dextran using sucrose or the synthesis of oligosaccharides when acceptor molecules like maltose are present. The DSRB742 gene (dsrB742) was cloned and the properties were characterized. In order to identify critical amino acid residues the DSRB742 amino acid sequence was aligned with glucosyltransferase sequences and three amino acid residues reported as sucrose binding amino acids in Streptococcus glucosyltransferases were selected for site-directed mutagenesis experiments. Asp-533 Asp- 536 and His-643 were independently replaced with Ala or Asn. D533A and D536A dextransucrases showed reduced dextran synthesis activities 2.3% and 40.8% of DSRB742 dextransucrase respectively and D533N D536N H643A and H643N dextransucrases showed complete suppression of dextran synthesis activities altogether. Additionally D536N dextransucrase showed complete suppression of oligosaccharide synthesis activities. However modifications at Asp-533 or at His-643 retained acceptor reaction activities in the range of 8.4% to 21.3% of DSRB742 acceptor reaction activity. Thus at least two carboxyl groups of Asp-533 and Asp-536 and His-643 as a proton donor are essential for the catalysis process.
|
|
|
KEYWORD
|
|
|
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|