A'Bháird N N, Ramsay R R
Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Biochem J. 1992 Sep 1;286 ( Pt 2)(Pt 2):637-40. doi: 10.1042/bj2860637.
Although the malonyl-CoA sensitivity of peroxisomal carnitine octanoyltransferase (COT) is reportedly lost on solubilization, we show that malonyl-CoA does inhibit the purified enzyme. Assay conditions such as buffer composition, pH, acyl-CoA substrate and the presence or absence of BSA can affect the observed inhibition. When assayed in the absence of BSA, COT shows simple competitive inhibition by malonyl-CoA. The Ki value for inhibition of purified COT is high (106 microM) compared with physiological concentrations (1-6 microM) and other short-chain acyl-CoA esters inhibit COT to the same degree. However, when COT is assayed in intact peroxisomes, the Ki for malonyl-CoA is almost 20-fold lower than found with the purified enzyme, whereas inhibition by other short-chain acyl-CoA esters does not change significantly. Several features of the inhibition of peroxisomal COT, including the specificity of malonyl-CoA over other short-chain acyl-CoA esters, resemble those of carnitine palmitoyltransferase (CPT)-I, suggesting that the regulation of COT and CPT-I in parallel may be necessary for the control of cellular fatty acid metabolism.
尽管据报道，过氧化物酶体肉碱辛酰转移酶（COT）的丙二酰辅酶A敏感性在溶解后丧失，但我们发现丙二酰辅酶A确实能抑制纯化后的该酶。诸如缓冲液组成、pH值、酰基辅酶A底物以及牛血清白蛋白（BSA）的存在与否等测定条件，都会影响所观察到的抑制作用。在不存在BSA的情况下进行测定时，COT表现出丙二酰辅酶A的简单竞争性抑制作用。与生理浓度（1 - 6 microM）相比，纯化后的COT的抑制常数（Ki）值较高（106 microM），并且其他短链酰基辅酶A酯对COT的抑制程度相同。然而，当在完整的过氧化物酶体中对COT进行测定时，丙二酰辅酶A的Ki值比纯化后的酶低近20倍，而其他短链酰基辅酶A酯的抑制作用没有显著变化。过氧化物酶体COT抑制作用的几个特征，包括丙二酰辅酶A相对于其他短链酰基辅酶A酯的特异性，类似于肉碱棕榈酰转移酶（CPT）-I的特征，这表明平行调节COT和CPT-I对于控制细胞脂肪酸代谢可能是必要的。
