GLYCEROL AS AN ENZYME-STABILIZING AGENT - EFFECTS ON ALDEHYDE DEHYDROGENASE
GLYCEROL AS AN ENZYME-STABILIZING AGENT - EFFECTS ON ALDEHYDE DEHYDROGENASE
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DOI：
10.1073/pnas.69.9.2373
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发表时间：
1972-01-01
影响因子：
11.1
通讯作者：
JAKOBY, WB
中科院分区：
文献类型：
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作者：
BRADBURY, SL;JAKOBY, WB
来自酵母的钾依赖性醛脱氢酶（EC 1.2.1.3）通过向水性缓冲液中加入高浓度的甘油或其它多元醇而显著改变。几条证据表明，活性位点附近的三维结构参与：（i）酶在2 ℃下储存或反复冻融时的稳定性取决于至少30%（v/v）甘油的存在。(ii)在相同的溶剂中，theKm值为DPN和苯甲醛的结合常数分别降低3和10倍，相比，在完全水性介质中获得的这些底物的值。(iii)在甘油中不再观察到三价砷对DPN的竞争性抑制;亚砷酸盐和Mapharsen的抑制作用变得混合，Ki值分别增加了5倍和50倍。(iv)必需的巯基，这是很容易羧甲基化在水性缓冲液中，是不容易获得的甘油或mannitol.The数据是一致的，在拓扑结构的变化引起的多元醇，其中附近的DPN结合位点的巯基被置换到一个更受保护的环境，在那里他们的反应性降低。由于这种醇的稳定作用是经常遇到的，这些结果可能有其他酶的应用。
The potassium-dependent aldehyde dehydrogenase (EC 1.2.1.3), from yeast is markedly altered by the addition of high concentrations of glycerol or other polyhydric alcohols to aqueous buffers. Several lines of evidence suggest that the three-dimensional structure near the active site is involved: (i) The stability of the enzyme when stored at 2°, or when subjected to repeated freezing and thawing, depends upon the presence of at least 30% (v/v) glycerol. (ii) In the same solvent, theKmvalue for DPN and the binding constant for benzaldehyde decrease by 3- and 10-fold, respectively, compared with the values obtained for these substrates in fully aqueous media. (iii) Competitive inhibition by trivalent arsenicals with respect to DPN is no longer observed in glycerol; the inhibition becomes mixed and theKivalues increase by 5- and 50-fold, respectively, with arsenite and Mapharsen. (iv) Essential sulfhydryl groups, which are easily carboxymethylated in aqueous buffers, are not readily available in either glycerol or mannitol.The data are consistent with a change in topography induced by polyhydric alcohols in which sulfhydryl groups near the DPN-binding site are displaced to a more protected environment, where their reactivity is reduced. Since the stabilizing effects of such alcohols are frequently encountered, these results may have application to other enzymes.