THE PHOSPHORYLATION SITE OF SNAKE MUSCLE FRUCTOSE 1,6-BISPHOSPHATASE AND ITS PHOSPHORYLATION CONDITIONS

From the tryptic digests of phosphorylated snake muscle FruP_(2ase), a phosphoryl peptide has beenisolated, its amino acid sequence was Gly-Ala-Gly-Ser-Arg and the phosphorylation site was consideredto be on the serine residue. Effectors of the enzyme such as FruP_2, F6P and AMP did not affect thephosphorylation. The effect of pH on phosphorylation was consistent with that on the activity of theenzyme. The activity of phosphorylated enzyme was slightly lower than that of the native enzyme, thisdifference in activities between the two forms of the enzyme increased with decreasing the substrateconcentration. Results further support that a phosphorylated intermediate is involved in the catalytic reac-tion of FruP_(2ase).     

蛇肌果糖1,6-二磷酸酯酶的磷酰化部位与磷酰化条件

本文报道了从磷酰化的果糖1,6-二磷酸酯酶的胰蛋白酶水解物中分离得到了一个磷酰化的肽段及该肽段的氨基酸排列顺序和磷酰化的部位是在丝氨酸残基上。果糖1,6-二磷酸,果糖6磷酸和腺嘌呤核苷-磷酸等效应剂不影响该酶被无机正磷酸磷酰化。pH对磷酰化程度的影响和对酶活力的影响一致。磷酰化酶的活力比非磷酰化状态的酶活力稍低,活力的差别随着底物浓度的减小而增大。磷酰化的酶与Mg~(2+)和果糖1,6-二磷酸一起保温能脱磷酰化,本文结果进一步支持了蛇肌果糖1,6-二磷酸酯酶在催化果糖1,6-二磷酸水解的过程中,包含了形成磷酰化酶中间物这一过程。     

蛇肌果糖1,6-二磷酸酯酶的逆反应中的磷酰化作用

本文证明在蛇肌果糖1,6-二磷酸酯酶的反应体系中,若有非水磷酸受体存在,其底物果糖1,6-二磷酸的1位磷酰基能转移到这些受体上形成有机磷化合物,导致F6P的释放速度快于无机磷的释放。用~(32)P-无机正磷酸和蛇肌酶一起保温,可分离出被无机磷共价磷酰化的酶中间物。这个磷酰化中间物可能和酶的正反应催化过程有关。     

PHOSPHORYLATED REACTION OF SNAKE MUSCLE FRUCTOSE 1,6-BISPHOSPHATASE IN ITS REVERSE REACTION

A phosphoryl enzyme intermediate was observed in the hydrolysis of FruP_2 catalyzed by snake muscleFruP_(2ase) based on the fact that the formation rate of F6P is faster than that of inorganic phosphatewhen the reaction mixture contains a phosphoryl acceptor other than water. A covalently bound phosphateon the enzyme was isolated from the reverse reaction. This intermediate may relate to the process of theforward catalytic reaction by the enzyme.