The molecular mechanism of beta-amylase inhibition against starch phosphorylase.

• Main Authors: Chen, Shih Ying, 陳師瑩700
• Other Authors: Su Jong-Ching, Lee Ping-Du
• Format: Others
• Language: zh-TW
• Published: 1997
• Online Access: http://ndltd.ncl.edu.tw/handle/30216219892791325079

博士 === 國立臺灣大學 === 農業化學系 === 85 === ABSTRACT The proteinaceous noncompetitive inhibitor of starch phosphorylase (SP)isolated from the root of sweet potato (Ipomoea batatas [L.] Lam.) has been identified as a beta- amylase (BA). We investigated three possible molecular mechanisms of BA inhibition against SP, namely an end-product inhibition of SPor BA, the modification of kinetic properties on binding or interaction of BA andSP, and the depletion of starch primer by the amylolytic action of BA. For assessing these mechanisms, dynamic light scattering (DLS), various designs ofenzyme kinetics, preparation of intermediate end-products (ILDs), electrophoreticmeasurements,carbohydrates analyzer (DIONEX) and GCG-program analysis were performed. The data indicated that the inhibitory effect of BA toward SPwas due to neither deprivation of starch nor end-productinhibition. Although no direct binding of BA and SP was detected, theSP- glucan-BA interaction was implicated according to an enzyme kinetics theoryand partially DLS data. Thismechanism could explain all phenomena associatedwith the inhibition. Different types of glucans as categorized according to branched or linear,and long or short average chain length that can be transcribed in term ofnon-reducing end concentration showed different patterns of effect on theinhibition kinetics. The inhibition of SP by BA is seemingly a universalphenomenon; BA from different plant sources could cross inhibit SP fromdifferent sources in a noncompetitive mode. No such inhibition could beobserved with other classes of amylolytic enzymes, such as alpha-amylase andamyloglucosidase. Advanced DLS, sedimentation and starch-agarose electrophoresis measurementsaimed at providing direct evidence for the presence of ternary complex so far have yielded no clear cut data due to either self aggregation through the glucan bridge, or the very weak force of interaction. However,the interaction of BA inhibition against SP is an inevitable outcome. Accordingto the results of enzyme kinetics, advanced DLS, sedimentation and agaroseelectrophoresis measurements, X-ray crystallographic, subsite theory and takinginto consideration the structural characteristics of glucans, we draw theschematic view of the interaction of BA inhibition against SP as in conclusion.