Korangy F, Julin D A
Department of Chemistry and Biochemistry, University of Maryland, College Park 20742.
J Biol Chem. 1992 Jan 25;267(3):1727-32.
The RecD subunit of the RecBCD enzyme from Escherichia coli contains an amino acid sequence common to many enzymes which bind ATP or GTP (Gly-X-X-Gly-X-Gly-Lys-Thr). We have changed the conserved lysine residue (amino acid number 177) in the RecD protein to glutamine to investigate the role of RecD, and ATP-binding to RecD, in the enzymatic activities of RecBCD. The mutant RecD protein assembles with the RecB and RecC subunits and the mutant enzyme, designated RecBCD-K177Q, can be purified in the same way as the wild-type RecBCD enzyme. The mutant RecD subunit in RecBCD-K177Q is photolabeled to a lesser extent by the ATP analogue 8-azido-adenosine-5'-triphosphate than is the wild-type RecD subunit in RecBCD, suggesting that the mutation has reduced the affinity of RecD for ATP.
来自大肠杆菌的RecBCD酶的RecD亚基含有许多结合ATP或GTP的酶所共有的氨基酸序列（甘氨酸- X - X - 甘氨酸- X - 甘氨酸- 赖氨酸- 苏氨酸）。我们将RecD蛋白中保守的赖氨酸残基（第177位氨基酸）替换为谷氨酰胺，以研究RecD以及RecD与ATP的结合在RecBCD酶活性中的作用。突变型RecD蛋白与RecB和RecC亚基组装在一起，这种突变酶命名为RecBCD - K177Q，其纯化方式与野生型RecBCD酶相同。与RecBCD中的野生型RecD亚基相比，RecBCD - K177Q中的突变型RecD亚基被ATP类似物8 - 叠氮基腺苷- 5'- 三磷酸光标记的程度较低，这表明该突变降低了RecD对ATP的亲和力。
