Flournoy D S, Frey P A
Biochemistry. 1986 Oct 7;25(20):6036-43. doi: 10.1021/bi00368a030.
The pyruvate dehydrogenase component (E1) of the pyruvate dehydrogenase complex catalyzes the decomposition of 3-fluoropyruvate to CO2, fluoride anion, and acetate. Acetylthiamin pyrophosphate (acetyl-TPP) is an intermediate in this reaction. Incubation of the pyruvate dehydrogenase complex with 3-fluoro[1,2-14C]pyruvate, TPP, coenzyme A (CoASH), and either NADH or pyruvate as reducing systems leads to the formation of [14C]acetyl-CoA. In this reaction the acetyl group of acetyl-TPP is partitioned by transfer to both CoASH (87 +/- 2%) and water (13 +/- 2%). When the E1 component is incubated with 3-fluoro[1,2-14C]pyruvate, TPP, and dihydrolipoamide, [14C]acetyldihydrolipoamide is produced. The formation of [14C]acetyldihydrolipoamide was examined as a function of dihydrolipoamide concentration (0.25-16 mM). A plot of the extent of acetyl group partitioning to dihydrolipoamide as a function of 1/[dihydrolipoamide] showed 95 +/- 2% acetyl group transfer to dihydrolipoamide when dihydrolipoamide concentration was extrapolated to infinity. It is concluded that acetyl-TPP is chemically competent as an intermediate for the pyruvate dehydrogenase complex catalyzed oxidative decarboxylation of pyruvate.
丙酮酸脱氢酶复合体的丙酮酸脱氢酶组分（E1）催化3-氟丙酮酸分解为二氧化碳、氟离子和乙酸盐。乙酰硫胺素焦磷酸（乙酰-TPP）是该反应的中间体。将丙酮酸脱氢酶复合体与3-氟[1,2-¹⁴C]丙酮酸、TPP、辅酶A（CoASH）以及作为还原系统的NADH或丙酮酸一起温育，会导致[¹⁴C]乙酰辅酶A的形成。在该反应中，乙酰-TPP的乙酰基通过转移至CoASH（87±2%）和水（13±2%）进行分配。当E1组分与3-氟[1,2-¹⁴C]丙酮酸、TPP和二氢硫辛酰胺一起温育时，会产生[¹⁴C]乙酰二氢硫辛酰胺。研究了[¹⁴C]乙酰二氢硫辛酰胺的形成与二氢硫辛酰胺浓度（0.25 - 16 mM）的关系。将乙酰基转移至二氢硫辛酰胺的程度作为1/[二氢硫辛酰胺]的函数作图，当二氢硫辛酰胺浓度外推至无穷大时，显示95±2%的乙酰基转移至二氢硫辛酰胺。结论是，乙酰-TPP在化学上能够作为丙酮酸脱氢酶复合体催化丙酮酸氧化脱羧反应的中间体。
