Watmough N J, Turnbull D M, Sherratt H S, Bartlett K
Department of Neurology, University of Newcastle upon Tyne, U.K.
Biochem J. 1989 Aug 15;262(1):261-9. doi: 10.1042/bj2620261.
The quantitative isolation of acyl-CoA esters of chain length C2-C17 from mitochondrial incubations and their analysis by reverse-phase radio-h.p.l.c. is described. Photodiode-array detection was used to characterize 2-enoyl-CoA esters. The chromatographic behaviour of all 27 intermediates of the beta-oxidation of hexadecanoyl-CoA is documented. Only C16, C14 and C12 intermediates were detected in uncoupled mitochondria oxidizing [U-14C]hexadecanoyl-CoA in the presence of fluorocitrate and carnitine, providing evidence for some organization of the enzymes of beta-oxidation [Garland, Shepherd & Yates (1965) Biochem. J. 97, 587-594; Sumegi & Srere (1984) J. Biol. Chem. 259, 8748-8752]. Rotenone increased concentrations of 3-hydroxyacyl-CoA and 2-enoyl-CoA esters and inhibited flux. These experiments provide the first direct unambiguous measurements of acyl-CoA esters in intact respiring rat liver mitochondrial fractions.
本文描述了从线粒体培养液中定量分离链长为C2 - C17的酰基辅酶A酯，并通过反相放射性高效液相色谱法对其进行分析的方法。利用光电二极管阵列检测对2-烯酰基辅酶A酯进行表征。记录了十六烷酰辅酶Aβ-氧化的所有27种中间体的色谱行为。在存在氟柠檬酸和肉碱的情况下，对氧化[U-14C]十六烷酰辅酶A的解偶联线粒体进行检测，仅检测到C16、C14和C12中间体，这为β-氧化酶的某种组织形式提供了证据[加兰、谢泼德和耶茨（1965年）《生物化学杂志》97卷，587 - 594页；苏梅吉和斯特雷（1984年）《生物化学杂志》259卷，8748 - 8752页]。鱼藤酮增加了3-羟酰基辅酶A和2-烯酰基辅酶A酯的浓度并抑制了通量。这些实验首次对完整呼吸的大鼠肝线粒体组分中的酰基辅酶A酯进行了直接明确的测量。
