Aldehyde Dehydrogenases in Rat Brain. Subcellular Distribution and Properties
Aldehyde Dehydrogenases in Rat Brain. Subcellular Distribution and Properties
复制标题
大鼠脑中的醛脱氢酶。
作者：
H. Pettersson;O. Tottmar
动力学研究表明，大鼠脑中存在几种形式的NAD依赖性醛脱氢酶（ALDH）。亚细胞分布研究表明，低- Km和高- Km的乙醛活性以及底物特异性酶琥珀酸半醛脱氢酶主要位于线粒体室。低Km活性也存在于细胞质中（<20%）。在以乙醛为底物的匀浆中，低Km活性仅为总活性的10-15%。用乙醛（0.2和2000 μm）和3,4‐二羟基苯乙醛（DOPAL）（0.3和31 μm）分别获得2个Km值，用琥珀酸半醛（5 μm）获得1个Km值。纯化后的线粒体经超声处理后，主要的醛脱氢酶与乙醛、DOPAL和琥珀酸半醛具有活性，但外膜标记酶（单胺氧化酶，MAO）的活性很小。在低渗缓冲液中，只有少量的ALDH活性从肿胀的线粒体中释放出来，而膜间隙标记酶（腺苷酸激酶）的主要部分被释放出来。这些结果表明ALDH与乙醛、DOPAL和琥珀酸半醛的活性位于基质室中。磷酸盐缓冲液中的Mg2+和Ca2+显著刺激了乙醛和DOPAL的低Km活性，而不是高Km活性和琥珀酸半醛脱氢酶。在焦磷酸盐缓冲液中，低- Km和高- Km对乙醛的活性表现出不同的pH最佳值。
Abstract: Kinetic studies suggested the presence of several forms of NAD‐dependent aldehyde dehydrogenase (ALDH) in rat brain. A subcellular distribution study showed that low‐ and high‐Km activities with acetaldehyde as well as the substrate‐specific enzyme succinate semialdehyde dehydrogenase were located mainly in the mitochondrial compartment. The low‐Km activity was also present in the cytosol (<20%). The low‐Km activity in the homogenate was only 10–15% of the total activity with acetaldehyde as the substrate. Two Km values were obtained with both acetaldehyde (0.2 and 2000 μm) and 3,4‐dihydroxyphenylacetaldehyde (DOPAL) (0.3 and 31 μm), and one Km value with succinate semialdehyde (5 μm). The main part of the aldehyde dehydrogenase activities with acetaldehyde, DOPAL, and succinate semialdehyde, but only little activity of the marker enzyme for the outer membrane (monoamine oxidase, MAO), was released from a purified mitochondrial fraction subjected to sonication. Only small amounts of the ALDH activities were released from mitochondria subjected to swelling in a hypotonic buffer, whereas the main part of the marker enzyme for the intermembrane space (adenylate kinase) was released. These results indicate that the ALDH activities with acetaldehyde, DOPAL and succinate semialdehyde are located in the matrix compartment. The low‐Km activity with acetaldehyde and DOPAL, but not the high‐Km activities and succinate semialdehyde dehydrogenase, was markedly stimulated by Mg2+ and Ca2+ in phosphate buffer. The low‐ and high‐Km activities with acetaldehyde showed different pH optima in pyrophosphate buffer.