Crystal Structure and Mutational Analysis of Isomalto-dextranase, a Member of Glycoside Hydrolase Family 27.

ABSTRACT: Arthrobacter globiformis T6 isomalto-dextranase (AgIMD) is an enzyme that liberates isomaltose from the non-reducing end of a polymer of glucose, dextran. AgIMD is classified as a member of the glycoside hydrolase family (GH) 27, which comprises mainly ?-galactosidases and ?-N-acetylgalactosaminidases, whereas AgIMD does not show ?-galactosidase or ?-N-acetylgalactosaminidase activities. Here, we determined the crystal structure of AgIMD. AgIMD consists of the following three domains: A, C, and D. Domains A and C are identified as a (?/?)8-barrel catalytic domain and an antiparallel ?-structure, respectively, both of which are commonly found in GH27 enzymes. However, domain A of AgIMD has subdomain B, loop-1, and loop-2, all of which are not found in GH27 human ?-galactosidase. AgIMD in a complex with trisaccharide panose shows that Asp-207, a residue in loop-1, is involved in subsite +1. Kinetic parameters of the wild-type and mutant enzymes for the small synthetic saccharide p-nitrophenyl ?-isomaltoside and the polysaccharide dextran were compared, showing that Asp-207 is important for the catalysis of dextran. Domain D is classified as carbohydrate-binding module (CBM) 35, and an isomaltose molecule is seen in this domain in the AgIMD-isomaltose complex. Domain D is highly homologous to CBM35 domains found in GH31 and GH66 enzymes. The results here indicate that some features found in GH13, -31, and -66 enzymes, such as subdomain B, residues at the subsite +1, and the CBM35 domain, are also observed in the GH27 enzyme AgIMD and thus provide insights into the evolutionary relationships among GH13, -27, -31, -36, and -66 enzymes.

SUBMITTER: Okazawa Y 

PROVIDER: S-EPMC4646281 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature