Harrison E H, Walusimbi-Kisitu M
Department of Biological Chemistry, School of Medicine, Wright State University, Dayton, Ohio 45435.
Am J Physiol. 1988 Sep;255(3 Pt 2):H441-5. doi: 10.1152/ajpheart.1988.255.3.H441.
The properties and subcellular localization of fatty acyl-CoA oxidase (FAO) were studied in rat heart homogenates. After differential centrifugation, FAO was sedimentable and enriched in a "light-mitochondrial" fraction. FAO had a pH optimum of 8-9. Among straight-chain, saturated fatty acyl-CoAs, the enzyme showed a marked preference for medium chain substrates (C12 greater than C10 = C8 greater than C16 = C14 greater than C6) over a concentration range up to 100 microM. No activity was observed with C4-CoA. The apparent Michaelis constant (Km) for C12-CoA was 5-10 microM. After removal of nuclei by low-speed centrifugation, combined subcellular particle preparations were obtained by high-speed centrifugation and layered on linear density gradients of metrizamide. After density equilibration, FAO showed a symmetric distribution centered at p = 1.16-1.18, like that of the enzyme catalase, a marker for microperoxisomes. In contrast, enzyme markers for mitochondria, lysosomes, sarcolemma, and sarcoplasmic reticulum were recovered in low-density regions of the gradient. These results provide a direct demonstration of fatty acyl-CoA oxidase in cardiac tissue and its association with microperoxisomes.
在大鼠心脏匀浆中研究了脂肪酰基辅酶A氧化酶（FAO）的性质和亚细胞定位。差速离心后，FAO可沉淀并富集于“轻线粒体”组分中。FAO的最适pH为8 - 9。在直链饱和脂肪酰基辅酶A中，该酶在高达100微摩尔的浓度范围内，对中链底物（C12大于C10 = C8大于C16 = C14大于C6）表现出明显的偏好。未观察到C4 - 辅酶A的活性。C12 - 辅酶A的表观米氏常数（Km）为5 - 10微摩尔。通过低速离心去除细胞核后，通过高速离心获得合并的亚细胞颗粒制剂，并将其铺在甲泛葡胺的线性密度梯度上。密度平衡后，FAO呈现出以p = 1.16 - 1.18为中心的对称分布，类似于微过氧化物酶体标记物过氧化氢酶的分布。相比之下，线粒体、溶酶体、肌膜和肌浆网的酶标记物在梯度的低密度区域中回收。这些结果直接证明了心脏组织中存在脂肪酰基辅酶A氧化酶及其与微过氧化物酶体的关联。
