Cloning and characterization of a dextranase gene from Lipomyces starkeyi and its expression in Saccharomyces cerevisiae.
A dextranase-encoding cDNA from L. starkeyi KSM22 was isolated and characterized. The 2052 bp cDNA fragment (lsd1) harbouring the dextranase gene exhibited one open reading frame (ORF) composed of 1824 bp flanked by a 41 bp 5'-UTR and a 184 bp 3'-UTR, including a 27 bp poly(A) tail. The lsd1 gene contains no introns. The open reading frame encodes a 608 amino acid polypeptide (LSD1) with a 67.6 kDa predicted molecular mass. There was a 77% deduced amino acid sequence identity between the LSD1 dextranase and the dextranase from Penicillium minioluteum. The primary structure of LSD1 dextranase exhibits distant similarity with the enzymes of the glycosyl hydrolase family 49 that comprises Penicillium dextranase. The optimum pH of LSD1 was 6.0 and the optimum temperature was 37 degrees C. LSD1 dextranase activity was substantially abolished by exposure to 1 mM Hg2+, Ag3+ and Mn2+. LSD1 exhibited high hydrolysing activity towards dextran (100%), soluble starch (22%) and mutan (8%).
Kang
Hee-Kyoung
HK
Laboratory of Functional Carbohydrate Enzymes and Microbial Genomics, Institute of Bioindustrial Technology, Chonnam National University, Gwang-Ju 500-757, South Korea.
Kim
Seung Heuk
SH
Park
Ji-Young
JY
Jin
Xing-Ji
XJ
Oh
Deok-Kun
DK
Kang
Seong Soo
SS
Kim
Doman
D
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003910:Dextranase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D004718:Saccharomycetales / Q000201:enzymology* / Q000235:genetics; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature
D011994:Recombinant Proteins; D003910:Dextranase