Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system: stereospecificity of proton transfer in the phosphorylation of enzyme I from (Z)-phosphoenolbutyrate.
Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system: stereospecificity of proton transfer in the phosphorylation of enzyme I from (Z)-phosphoenolbutyrate.
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大肠杆菌磷酸烯醇丙酮酸依赖性磷酸转移酶系统：(Z)-磷酸烯醇丁酸酶 I 磷酸化过程中质子转移的立体特异性。
DOI：
10.1021/bi00281a010
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发表时间：
1983
期刊：
影响因子：
2.9
通讯作者：
G. Robillard
中科院分区：
文献类型：
--
作者：
H. Hoving;T. Nowak;G. Robillard
建立了磷酸烯醇丁酸酯与酶Ⅰ反应中质子转移的立体化学。在磷酸烯醇丙酮酸类似物的纯Z异构体与酶I反应以产生磷酸酶I和2-氧代丁酸酯期间，底物在2 re，3si面的C-3处质子化。这种立体特异性是针对质子转移到（Z）-磷酸[3-D]烯醇丁酸酯和氘转移到（Z）-磷酸[3-H]烯醇丁酸酯而建立的。磷酸烯醇丁酸酯的E异构体不是酶I的底物。因此，磷酸酶I与2-氧代丁酸酯的反应仅产生磷酸烯醇丁酸酯的Z异构体，并且仅提取2-氧代丁酸酯的C-3处的pro-S质子。在该反应中的动力学H/D同位素效应为6.8，表明了质子转移步骤的限速性质。利用丙酮酸激酶催化的烯醇化反应对2-氧代[3（R）-H，D]丁酸酯和2-氧代-[3（S）-H，D]丁酸酯进行了立体化学分析。这种酶促烯醇化反应，以磷酸盐作为辅因子，在中性pH下快速，并且是高度立体特异性的反应：仅2-氧代丁酸酯的C-3处的pro-R质子与溶剂交换。该反应还用于产生2-氧代[3-H，D]丁酸酯的纯3R和3S对映异构体。从甲基质子核磁共振信号的精细结构检测2-氧代丁酸酯的C-3处的质子化/氘化程度。
The stereochemistry of the proton transfer in the reaction of phosphoenolbutyrate with enzyme I has been established. During the reaction of the pure Z isomer of this analogue of phosphoenolpyruvate with enzyme I, to yield phosphoenzyme I and 2-oxobutyrate, the substrate is protonated at C-3 from the 2re,3si face. This stereospecificity was established for the transfer of a proton to (Z)-phospho[3-D]enolbutyrate and for the transfer of a deuteron to (Z)-phospho[3-H]enolbutyrate. The E isomer of phosphoenolbutyrate is not a substrate for enzyme I. Accordingly, the reaction of phosphoenzyme I with 2-oxobutyrate yields exclusively the Z isomer of phosphoenolbutyrate, and only the pro-S proton at C-3 of 2-oxobutyrate is abstracted. A kinetic H/D isotope effect of 6.8 in this reaction demonstrates the rate-limiting nature of the proton-transfer step. The stereochemical analysis of 2-oxo[3(R)-H,D]butyrate and of 2-oxo-[3(S)-H,D]butyrate was carried out by using the pyruvate kinase catalyzed enolization of this compound. This enzymatic enolization, with phosphate as a cofactor, is rapid at neutral pH and is a highly stereospecific reaction: only the pro-R proton at C-3 of 2-oxobutyrate is exchanged with solvent. This reaction was also used to generate the pure 3R and 3S enantiomers of 2-oxo[3-H,D]butyrate. The degree of protonation/deuteration at C-3 of 2-oxobutyrate was detected from the fine structure of the methyl proton nuclear magnetic resonance signal.
由细菌磷酸烯醇丙酮酸的酶 I 催化的磷酸基交换反应：糖磷酸转移酶系统。
DOI：
--
发表时间：
1980
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
SaierJr,MH;Schmidt,MR;Lin,P
通讯作者：
Lin,P