Hexose phosphate binding sites of fructose 6-phosphate,2-kinase:fructose 2,6-bisphosphatase.
Hexose phosphate binding sites of fructose 6-phosphate,2-kinase:fructose 2,6-bisphosphatase.
复制标题
果糖 6-磷酸,2-激酶:果糖 2,6-二磷酸酶的己糖磷酸结合位点。
DOI：
10.1021/bi00019a018
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发表时间：
1995
期刊：
影响因子：
2.9
通讯作者：
Uyeda,K
中科院分区：
文献类型：
--
作者：
Tsujikawa,T;Watanabe,F;Uyeda,K
Revised Manuscript Received January 31, 1995® abstract: A previous chemical modification study [Kitamura et al.(1989) J. Biol. Chem. 264, 6344—6438] has shown that A-bromoacetylethanolamine phosphate labeled specifically Cys 107 of rat liver Fru 6-P, 2-kinase: Fru 2, 6-Pase and the corresponding Cys of the bovine heart enzyme, leading to inactivation of kinase activity. Since Fru 6-P provided protection against the inactivation, this region of the enzyme was thought to be a Fru 6-P binding site of the kinase enzyme. To examine this possibility, oligonucleotidedirected mutagenesis has been used to alter several residues in expressedrat testis Fru 6-P, 2-kinase: Fru 2.6-Pase. The change of Lys 100, Lysl03, and Aspll2 caused at most a 2-fold increase in KmF6P and a 2—3-fold increase in ATmATP, suggesting that these residues are not involved in the directbinding of Fru 6-P. However, change of Argl02 to Leu and to Lys resulted in a 325 x and 22 x, respectively, increase in KmF6P, and change of Argl02 to Glu resulted in nearly complete loss of the kinase activity. Change of Cys 105 to Ala or Ser increased KmF6p about 10 x. The Vmax of all these mutated enzymes except the one that changed Argl02 to Glu (R102E) was increased 10% to 85%. The kinetic parameters of Fru 2.6-Pase were not altered by these changes. R102E formed several polymeric forms of the enzyme, including a tetramer. Both R102E and an additional derivative that substituted Lys for Argl02 (R102K) were slightly more susceptible to guanidine inactivation than the wild-type enzyme. These results suggest that Argl02 is essential for binding the 6-phosphate of Fru 6-P and that Cys 105 also may play a role at the same site.Synthesis and degradation of Fru 2, 6-P2, the most potent activator of phosphofructokinase, are catalyzed by a bifunc-tional enzyme, Fm 6-P, 2-kinase: Fm 2, 6-bisphosphatase (Fm 6-P+ ATP= Fm 2, 6-P2+ ADP and Fra 2, 6-P2—Fra 6-P
