Konstantinidis A K, Marsden I, Sinnott M L
Department of Chemistry, University of Illinois, Chicago 60680.
Biochem J. 1993 May 1;291 ( Pt 3)(Pt 3):883-8. doi: 10.1042/bj2910883.
Cellobiohydrolase II hydrolyses alpha- and beta-D-cellobiosyl fluorides to alpha-cellobiose at comparable rates, according to Michaelis-Menten kinetics. The stereochemistry, absence of transfer products and strict hyperbolic kinetics of the hydrolysis of alpha-cellobiosyl fluoride suggest that the mechanism for the alpha-fluoride may be the enzymic counterpart of the SNi reaction observed in the trifluoroethanolysis of alpha-glucopyranosyl fluoride [Sinnott and Jencks (1980) J. Am. Chem. Soc. 102, 2026-2032]. The absolute factors by which this enzyme accelerates fluoride ion release are small and greater for the alpha-fluoride than for the beta, suggesting that its biological function may not be just glycoside hydrolysis. Cellobiohydrolase I hydrolyses only beta-cellobiosyl fluoride, which is, however, an approx. 1-3% contaminant in alpha-cellobiosyl fluoride as prepared and purified by conventional methods. Instrumental assays for the various components of the cellulase complex are discussed.
根据米氏动力学,纤维二糖水解酶II以可比的速率将α-和β-D-纤维二糖氟化物水解为α-纤维二糖。α-纤维二糖氟化物水解的立体化学、无转移产物以及严格的双曲线动力学表明,α-氟化物的水解机制可能是在α-吡喃葡萄糖基氟化物的三氟乙醇解中观察到的SNi反应的酶促对应物[Sinnott和Jencks(1980年)《美国化学会志》102,2026 - 2032]。该酶加速氟离子释放的绝对因子较小,且α-氟化物的大于β-氟化物的,这表明其生物学功能可能不仅仅是糖苷水解。纤维二糖水解酶I仅水解β-纤维二糖氟化物,然而,在用常规方法制备和纯化的α-纤维二糖氟化物中,它是约1 - 3%的污染物。讨论了纤维素酶复合物各种成分的仪器分析方法。