ROLE OF EFFECTOR BINDING IN ALLOSTERIC CONTROL OF RIBONUCLEOSIDE DIPHOSPHATE REDUCTASE
ROLE OF EFFECTOR BINDING IN ALLOSTERIC CONTROL OF RIBONUCLEOSIDE DIPHOSPHATE REDUCTASE
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DOI：
10.1016/0022-2836(69)90056-4
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发表时间：
1969-01-01
影响因子：
5.6
通讯作者：
REICHARD, P
中科院分区：
文献类型：
--
作者：
BROWN, NC;REICHARD, P
变构效应分子ATP、dATP、dGTP和dTTP调节大肠杆菌核糖核苷二磷酸还原酶的活性水平和底物专一性。活性酶由两个不同亚基组成的复合体，即蛋白质B1和B2。添加负性效应物，如dATP，会导致形成一种非活性形式，它可能是活性酶的二聚体。我们现在已经研究了效应核苷酸与B1和B2亚基的结合。亚基B2不与任何核苷酸结合，而亚基B1单独或与亚基B2结合，总共包含四个结合位点。这些可分为两类，每类包含两个地点。第一类(h-位点)与dATP有较高的亲和力，可与ATP、dATP、dGTP和dTTP结合；第二类(L-位点)与dATP亲和力较低，仅与dATP和dATP结合。在蛋白B2的存在下，dATP与L位点的结合比在分离的蛋白B1中要紧密得多。这提示了一种通过与dATP结合来产生失活形式的酶的机制。似乎L-位点主要参与酶的整体活性的调节，而效应器与H-位点的结合影响底物特异性。不同的核苷酸与H-和L-位点结合形成许多不同种类的蛋白质-效应复合体，每种复合体都与酶的特定功能状态有关。这表明，核糖核苷二磷酸还原酶的调控需要存在许多不同的构象状态。
The allosteric effectors, ATP, dATP, dGTP and dTTP, regulate both the level of activity and the substrate specificity of ribonucleoside diphosphate reductase fromEscherichia coli. The active enzyme consists of a complex of two non-identical subunits, proteins B1 and B2. Addition of negative effectors, such as dATP, leads to the formation of an inactive form which is probably a dimer of the active enzyme. We have now studied the binding of the effector nucleotides to the B1 and B2 subunits. Subunit B2 does not bind any of the nucleotides, while subunit B1, either alone or in combination with subunit B2, contains a total of four binding sites. These can be divided into two classes, each containing two sites. The first class (h-sites) has a high affinity for dATP and binds ATP, dATP, dGTP and dTTP; the second class (l-sites) has a lower affinity for dATP and binds only ATP and dATP. Binding of dATP to l-sites is considerably tighter in the presence of protein B2 than in the isolated protein B1. This suggests a mechanism for the generation of the inactive form of the enzyme through the binding of dATP. It appears that thel-sitesare primarily involved in the regulation of theover-all activityof the enzyme, while binding of effectors to theh-sitesinfluences thesubstrate-specificity. The binding of different nucleotides to h- and l-sites results in the formation of many distinct species of protein-effector complexes, each of which can be related to particular functional states of the enzyme. This suggests that the regulation of ribonucleoside diphosphate reductase requires the existence of many distinct conformational states.