THE FORMALDEHYDE DEHYDROGENASE OF RHODOCOCCUS-ERYTHROPOLIS, A TRIMERIC ENZYME REQUIRING A COFACTOR AND ACTIVE WITH ALCOHOLS
THE FORMALDEHYDE DEHYDROGENASE OF RHODOCOCCUS-ERYTHROPOLIS, A TRIMERIC ENZYME REQUIRING A COFACTOR AND ACTIVE WITH ALCOHOLS
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DOI：
10.1111/j.1432-1033.1985.tb08997.x
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发表时间：
1985-01-01
期刊：
影响因子：
--
通讯作者：
SAHM, H
中科院分区：
文献类型：
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作者：
EGGELING, L;SAHM, H
在含有甲基的化合物上生长的过程中，革兰氏阳性菌红种革兰氏菌的甲醛脱氢酶被诱导。采用亲和层析和渗透层析对甲醛脱氢酶进行了纯化。酶的等电点为4.7。测定了天然酶的分子量为130,000 g/mol。十二烷基硫酸钠凝胶电泳得到单个亚基，分子量为44,000 g/mol。这些结果，以及交联实验产生的单体，二聚体和三聚体带，与甲醛脱氢酶的三聚亚基结构一致。以甲醛为底物的活性需要低MW的热稳定辅因子。这种辅因子被发现是可氧化的，但只有在其还原形式下才有活性。制备电聚焦结果表明，辅因子是一种弱酸，其pK值约为。6.5. 该酶在不需要辅因子存在的情况下，对从乙醇到辛醇等一系列伯醇具有活性。一个没有甲醛脱氢酶活性的突变体在乙醇作为底物的生长中没有受到损害。这表明，醇类模拟甲醛脱氢酶的真正底物，它可能是辅因子的羟基甲基衍生物，由甲醛的加入而产生。
During growth on compounds containing methyl groups, a formaldehyde dehydrogenase is induced in the gram-positive bacteria, R. erythropolis. This formaldehyde dehydrogenase was purified to homogeneity using affinity chromatography and permeation chromatography. The isoelectric point of the enzyme was 4.7. The MW of the native enzyme was determined as 130,000 g/mol. Sodium dodecyl sulfate gel electrophoresis yielded a single subunit with a MW of 44,000 g/mol. These results, together with cross-linking experiments which yielded monomer, dimer and trimer bands, are consistent with a trimeric subunit structure of the formaldehyde dehydrogenase. A heat-stable cofactor of low MW was required for activity with formaldehyde as substrate. This cofactor was found to be oxidizable, but active only in its reduced form. Preparative electrofocusing revealed that the cofactor is a weak acid with a pK of .apprx. 6.5. The enzyme was active with the homologous series of the primary alcohols, ethanol up to octanol, without requiring the presence of the cofactor. A mutant without formaldehyde dehydrogenase activity was not impaired in its growth with ethanol as substrate. It is suggested that the alcohols mimic the true substrate of the formaldehyde dehydrogenase, which could be a hydroxymethyl derivative of the cofactor, resulting from the addition of formaldehyde.