S-Nitrosoglutathione is a substrate for rat alcohol dehydrogenase class III isoenzyme
S-Nitrosoglutathione is a substrate for rat alcohol dehydrogenase class III isoenzyme
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DOI：
10.1042/bj3310659
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发表时间：
1998-04-15
影响因子：
4.1
通讯作者：
Du Bois, GC
中科院分区：
文献类型：
--
作者：
Jensen, DE;Belka, GK;Du Bois, GC
从大鼠肝胞质溶胶中分离的酶（天然分子量78.3 kDa;多肽分子量42.5 kDa）能够催化S-亚硝基谷胱甘肽（GSNO）的NADH/NADPH依赖性降解。该活性利用每摩尔处理的GSNO 1摩尔的辅酶。分离的酶也具有醇脱氢酶（ADH）III类同工酶所特有的几个特征：它能够催化辛醇（对4-甲基吡唑的抑制不敏感）、甲基巴豆醇（通过添加的戊酸刺激）和12-羟基十二烷酸的NAD（+）依赖性氧化，以及辛醛的NADH/NADPH依赖性还原。甲醇和乙醇的氧化活性极低。该酶具有甲醛脱氢酶活性，因为它能够催化S-羟甲基谷胱甘肽的NAD（+）/NADP（+）依赖性氧化。用精氨酸特异性试剂苯乙二醛处理可防止甲基巴豆醇氧化的戊酸刺激，并显著降低对辛醇、12-羟基十二烷酸和S-羟甲基谷胱甘肽的酶活性;还检查了催化GSNO降解的能力。此外，有限的肽测序表明与已知的ADH III类同工酶序列100%一致。动力学研究表明，GSNO是这种酶的一个非常活跃的底物。S-亚硝基-IV-乙酰青霉胺和S-亚硝化人血清白蛋白不是底物;对S-亚硝化谷胱甘肽单乙酯和二乙酯的活性极小。产物分析表明，谷胱甘肽亚磺酰胺是酶促GSNO加工的主要稳定产物，GSSG和NH的产率较小; GSH、羟胺、亚硝酸盐、硝酸盐和一氧化氮积累最小。在反应混合物中包含GSH降低了假定的谷胱甘肽亚磺酰胺的产率，有利于GSSG和羟胺。
An enzyme isolated from rat liver cytosol (native molecular mass 78.3 kDa; polypeptide molecular mass 42.5 kDa) is capable of catalysing the NADH/NADPH-dependent degradation of S-nitrosoglutathione (GSNO). The activity utilizes 1 mol of coenzyme per mol of GSNO processed. The isolated enzyme has, as well, several characteristics that are unique to alcohol dehydrogenase (ADH) class III isoenzyme: it is capable of catalysing the NAD(+)-dependent oxidations of octanol (insensitive to inhibition by 4-methylpyrazole), methylcrotyl alcohol (stimulated by added pentanoate) and 12-hydroxydodecanoic acid, and also the NADH/NADPH-dependent reduction of octanal. Methanol and ethanol oxidation activity is minimal. The enzyme has formaldehyde dehydrogenase activity in that it is capable of catalysing the NAD(+)/NADP(+)-dependent oxidation of S-hydroxymethylglutathione. Treatment with the arginine-specific reagent phenylglyoxal prevents the pentanoate stimulation of methylcrotyl alcohol oxidation and markedly diminishes the enzymic activity towards octanol, 12-hydroxydodecanoic acid and S-hydroxymethylglutathione; the capacity to catalyse GSNO degradation is also checked. Additionally, limited peptide sequencing indicates 100 % correspondence with known ADH class III isoenzyme sequences. Kinetic studies demonstrate that GSNO is an exceptionally active substrate for this enzyme. S-Nitrosa-IV-acetylpenicillamine and S-nitrosated human serum albumin are not substrates; the activity towards S-nitrosated glutathione mono-and di-ethyl esters is minimal. Product analysis suggests that glutathione sulphinamide is the major stable product of enzymic GSNO processing, with minor yields of GSSG and NH,; GSH, hydroxylamine, nitrite, nitrate and nitric oxide accumulations are minimal. Inclusion of GSH in the reaction mix decreases the yield of the supposed glutathione sulphinamide in favor of GSSG and hydroxylamine.