SPATIAL ARRANGEMENT OF COENZYME AND SUBSTRATES BOUND TO L-3-HYDROXYACYL-COA DEHYDROGENASE AS STUDIED BY SPIN-LABELED ANALOGS OF NAD+ AND COA
SPATIAL ARRANGEMENT OF COENZYME AND SUBSTRATES BOUND TO L-3-HYDROXYACYL-COA DEHYDROGENASE AS STUDIED BY SPIN-LABELED ANALOGS OF NAD+ AND COA
复制标题
DOI：
10.1021/bi00225a007
复制
发表时间：
1991-03-19
期刊：
影响因子：
2.9
通讯作者：
TROMMER, WE
中科院分区：
文献类型：
--
作者：
HARTMANN, D;PHILIPP, R;TROMMER, WE
描述了s -乙酰乙酰辅酶a、s -乙酰乙酰泛茶氨酸和s -乙酰乙酰半胱胺的氮氧自旋标记衍生物的合成。这些化合物是l- 3-羟基酰基辅酶a脱氢酶的活性底物[(S)-3-羟基酰基辅酶a:NAD+氧化还原酶，EC 1.1.1.35]，其upsilon-max值为S-乙酰乙酰辅酶a本身的20%至70%。S-Acetoacetylpantetheine和s - acetoacetyl半胱胺与酶形成二元配合物，并表现出典型的固定化氮氧化物的ESR光谱。在自旋标记的泛酸中，自由基的流动性更强。当自旋标记的底物同时结合到这种二聚体酶的每个活性位点时，自旋-自旋相互作用区分了底物的两个可选方向[Birktoft， J. J., Holden， H. M., Hamlin， R., Xuong， N. H., & Banaszak， L. J. (1987) Proc. Natl.]学会科学。[j]。脂肪酸部分被认为位于两个结构域之间的间隙中，而CoA部分的大部分可能延伸到溶液中。NAD+在腺嘌呤环的N6处自旋标记，是l- 3-羟基酰基辅酶a脱氢酶的活性辅酶（60% upsilon-max）。该酶的配合物表现出高度固定化氮氧化物的典型ESR光谱。自旋标记的s -乙酰乙酰基泛茶氨酸的ESR谱变化揭示了辅酶NAD+的结合导致二元酶/底物复合物的构象变化。
The synthesis of nitroxide spin-labeled derivatives of S-acetoacetyl-CoA, S-acetoacetylpantetheine, and S-acetoacetylcysteamine is described. These compounds are active substrates of L-3-hydroxyacyl-CoA dehydrogenase [(S)-3-hydroxyacyl-CoA:NAD+ oxidoreductase, EC 1.1.1.35] exhibiting upsilon-max values from 20% to 70% of S-acetoacetyl-CoA itself. S-Acetoacetylpantetheine and S-acetoacetylcysteamine form binary complexes with the enzyme and exhibit ESR spectra typical for immobilized nitroxides. In the case of spin-labeled pantetheine, the radical is more mobile. When spin-labeled substrates are bound simultaneously to each active site of this dimeric enzyme, spin-spin interactions differentiate between two alternate orientations of the substrate [Birktoft, J. J., Holden, H. M., Hamlin, R., Xuong, N. H., & Banaszak, L. J. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 8262-8266]. The fatty acid moiety is thought to be located in a cleft between two domains whereas a large part of the CoA moiety probably extends into the solution. NAD+, spin-labeled at N6 of the adenine ring, is an active coenzyme of L-3-hydroxyacyl-CoA dehydrogenase (60% upsilon-max). Complexes with the enzyme exhibit ESR spectra typical of highly immobilized nitroxides. Binding of coenzyme NAD+ causes conformational changes of the binary enzyme/substrate complex as revealed by changes in the ESR spectrum of spin-labeled S-acetoacetylpantetheine.