Passonneau J V, Schwartz J P, Rottenberg D A
J Biol Chem. 1975 Mar 25;250(6):2287-92.
Both the I (independent of glucose 6-phosphate) and D (dependent on glucose 6-phosphate) forms of glycogen synthase (UDP-glucose:glycogen alpha-4-glucosyltransferase EG 2.4.1.11) have been partially purified from pig brain and the kinetic constants of the enzymes have been examined. The Km for UDP-glucose for the I form increased from 0.11 to 0.5 mM when the temperature was raised from 25 to 37 degrees. When glucose 6-phosphate was present, the Km for UDP-glucose was decreased to 0.03 and 0.08 mM at 25 and 37 degrees, respectively. The amount of glucose 6-phosphate required to produce half-maximal stimulation decreased with increasing UDP-glucose concentration at both temperatures but increased with increasing temperature. The Km for glucose 6-phosphate at 0.03 and 0.20 mM UDP-glucose was 0.13 and 0.10 mM, respectively, at 25 degrees. At 37 degrees and 0.125 and 4.0 mM UDP-glucose the Km for glucose 6-phosphate was 0.32 and 0.04 mM, respectively. The Km for UDP-glucose for the D form at 0.75, 2.0, and 10 mM glucose 6-phosphate was 0.71, 0.50, and 0.42 mM at 25 degrees. At higher temperatures the apparent affinity for the substrate was decreased; at 37 degrees, the Km for UDP-glucose at 0.75 and 2.0 nM glucose 6-phosphate was 5.75 and 1.42 mM, respectively. The requirement for glucose 6-phosphate was decreased when UDP-glucose concentrations were increased; at 0.5 and 5.0 mM UDP-glucose concentrations, the Km for glucose 6-phosphate was 22.7 and 1.82 mM at 25 degrees. As was the case with the I form, the apparent Km for glucose 6-phosphate increased at higher temperatures. At 37 degrees, the Km for glucose 6-phosphate at 0.5 and 5.0 mM UDP-glucose was 43.5 and 6.15 mM. The temperature coefficient for the maximum velocity was 10.1% per degree for synthase I and 8.5% per degree for synthase D between 25 and 37 degrees. The D form of synthase was calculated to be virtually inactive under normal physiological conditions with the substrate concentrations found in the brain. The enzymatic activity calculated for synthase I correlates well with the observed rate of incorporation of UDP-[U-14C]glucose into brain glycogen.
已从猪脑中部分纯化出糖原合酶（UDP - 葡萄糖：糖原α - 4 - 葡糖基转移酶，EC 2.4.1.11）的I（不依赖6 - 磷酸葡萄糖）和D（依赖6 - 磷酸葡萄糖）两种形式，并对这些酶的动力学常数进行了检测。I型糖原合酶对UDP - 葡萄糖的Km值在温度从25℃升高到37℃时，从0.11 mM增加到0.5 mM。当存在6 - 磷酸葡萄糖时，在25℃和37℃下，UDP - 葡萄糖的Km值分别降至0.03 mM和0.08 mM。在两个温度下，产生最大刺激一半所需的6 - 磷酸葡萄糖量随UDP - 葡萄糖浓度增加而减少，但随温度升高而增加。在25℃时，UDP - 葡萄糖浓度为0.03 mM和0.20 mM时，6 - 磷酸葡萄糖的Km值分别为0.13 mM和0.10 mM。在37℃、UDP - 葡萄糖浓度为0.125 mM和4.0 mM时，6 - 磷酸葡萄糖的Km值分别为0.32 mM和0.04 mM。在25℃时，D型糖原合酶在6 - 磷酸葡萄糖浓度为0.75 mM、2.0 mM和10 mM时，对UDP - 葡萄糖的Km值分别为0.71 mM、0.50 mM和0.42 mM。在较高温度下，对底物的表观亲和力降低；在37℃时，6 - 磷酸葡萄糖浓度为0.75 mM和2.0 mM时，D型糖原合酶对UDP - 葡萄糖的Km值分别为5.75 mM和1.42 mM。当UDP - 葡萄糖浓度增加时，对6 - 磷酸葡萄糖的需求降低；在25℃时，UDP - 葡萄糖浓度为0.5 mM和5.0 mM时，6 - 磷酸葡萄糖的Km值分别为22.7 mM和1.82 mM。与I型糖原合酶情况相同，在较高温度下，6 - 磷酸葡萄糖的表观Km值增加。在37℃时，UDP - 葡萄糖浓度为0.5 mM和5.0 mM时，6 - 磷酸葡萄糖的Km值分别为43.5 mM和6.15 mM。在25℃至37℃之间，合酶I的最大反应速度的温度系数为每度10.1%，合酶D为每度8.5%。计算得出，在大脑中发现的底物浓度下，D型合酶在正常生理条件下几乎无活性。计算得出的合酶I的酶活性与观察到的UDP - [U - 14C]葡萄糖掺入脑糖原的速率密切相关。
