Monday, August 2, 2010
Pacific Concourse (Hyatt Regency San Francisco)
Amylosucrase (ASase, EC 2. 4. 1. 4) is a glycosyltrasnferase that transfer an glucose from sucrose to the 4-positisions of the non-reducing end of an α-1→4 glucan chain of the acceptor molecule, thereby creating an amylose-like polymer. When sucrose is used as a sole substrate, ASase proceeds three-step catalytic reactions : 1) Fructose is released by forming glucosyl-enzyme intermediate (Total activity). 2) Hydrolysis reaction produces glucose by attack of water on the glycosyl-intermediate (Hydrolysis activity). 3) Transglycosylation was occurred if the water molecule is replaced by a competing acceptor such as glucose, maltose, and alcohols (Transglycosylation activity). ASase gene was cloned from Deinococcus geothermalis (dgas) and efficiently expressed in E. coli using glutathione S-transferase fusion system. The DGAS reaction kinetics using sucrose as substrate was performed at 30°C and 45°C to investigate into DGAS dual activities by temperature. The Linewear-Burk representation demonstrates that DGAS has favorite behavior between total activity, hydrolysis activity and transglycosylation activity by temperature. As reaction temperature increased, from 25°C to 45°C, ratio of hydrolysis activity also improved whereas ratio of transglycosylation activity was decreased in the presence of sucrose as sole substrate.