Ball K, Preiss J
Department of Biochemistry, Michigan State University, East Lansing 48824.
J Biol Chem. 1994 Oct 7;269(40):24706-11.
Pyridoxal-P, an analog of 3-phosphoglycerate, activates spinach leaf ADPglucose pyrophosphorylase. Reductive phosphopyridoxylation of the enzyme yields enzyme less dependent on the presence of activator for activity. Labeled pyridoxal-P is incorporated into both the 51- and 54-kDa subunits of the enzyme. The sequence of the single tryptic labeled peptide of the small subunit has been reported (Morell, M., Bloom, M., and Preiss, J. (1988) J. Biol. Chem. 263, 633-637). Three distinct labeled peptides are isolated from the tryptic digest of the large subunit. 3-Phosphoglycerate inhibits incorporation of pyridoxal-P by 80% into all three large subunit tryptic peptides, whereas inorganic phosphate, the allosteric inhibitor, inhibits incorporation into only one of those peptides. Comparison of the sequences of the labeled tryptic peptides with those of large and small subunits of the enzyme from other species indicate their positions in the primary structure of the spinach enzyme large subunit. The conservation in the amino acid sequences surrounding the lysyl residue in the small subunit and of the lysyl residue in the large subunit in higher plant and in cyanobacterial ADPglucose pyrophosphorylases, which are protected from phosphopyridoxylation by both allosteric effectors, suggests that these lysyl residues are involved in activator binding.
磷酸吡哆醛P是3-磷酸甘油酸的类似物，可激活菠菜叶ADP葡萄糖焦磷酸化酶。该酶的还原性磷酸吡哆醛化作用使酶在活性上对激活剂的存在依赖性降低。标记的磷酸吡哆醛P被掺入该酶的51kDa和54kDa亚基中。小亚基的单一胰蛋白酶标记肽的序列已有报道（莫雷尔，M.，布鲁姆，M.，和普赖斯，J.（1988年）《生物化学杂志》263卷，633 - 637页）。从大亚基的胰蛋白酶消化物中分离出三种不同的标记肽。3-磷酸甘油酸可抑制磷酸吡哆醛P向所有三种大亚基胰蛋白酶肽中的掺入达80%，而变构抑制剂无机磷酸盐仅抑制向其中一种肽中的掺入。将标记的胰蛋白酶肽的序列与来自其他物种的该酶大亚基和小亚基的序列进行比较，表明了它们在菠菜酶大亚基一级结构中的位置。高等植物和蓝细菌ADP葡萄糖焦磷酸化酶中小亚基中赖氨酸残基周围以及大亚基中赖氨酸残基周围氨基酸序列的保守性，这两种残基都受到变构效应物的保护而不被磷酸吡哆醛化，表明这些赖氨酸残基参与激活剂的结合。
