Khatri Yogan, Hannemann Frank, Girhard Marco, Kappl Reinhard, Même Aurélie, Ringle Michael, Janocha Simon, Leize-Wagner Emmanuelle, Urlacher Vlada B, Bernhardt Rita
Department of Biochemistry, Saarland University, Saarbrücken, Germany.
Biotechnol Appl Biochem. 2013 Jan-Feb;60(1):18-29. doi: 10.1002/bab.1087.
The members of the CYP109 family (CYP109C1, CYP109C2, and CYP109D1) from Sorangium cellulosum So ce56 are among the 21 P450 enzymes, of which only CYP109D1 and CYP264B1 have so far been functionally characterized. Here, we attempted to characterize two other P450s (CYP109C1 and CYP109C2) for the first time and compare their biochemical, biophysical, and functional properties to those of the fatty acid hydroxylating CYP109D1. Considering the physiological importance of fatty acids, we investigated saturated fatty acid binding and conversion for all members of the CYP109 family. The interaction between the CYP109 members and different autologous/heterologous redox partners was compared using Biacore measurements in which only CYP109D1 and bovine adrenodoxin (Adx) formed a complex. Surprisingly, this interaction was similarly efficient as the interaction of Adx with its mammalian redox partners. The in vitro reconstitution assays showed no activity when using CYP109C1, although substrate binding was demonstrated; also, there was subterminal hydroxylation of saturated fatty acids, when using CYP109C2 and CYP109D1, where CYP109D1 was a much more efficient fatty acid hydroxylase. Interestingly, the hydroxylation position moved inside the fatty acid chain when using long-chain fatty acids, thus producing possible precursors for physiologically important products.
来自纤维堆囊菌So ce56的CYP109家族成员（CYP109C1、CYP109C2和CYP109D1）属于21种细胞色素P450酶，其中到目前为止只有CYP109D1和CYP264B1的功能得到了表征。在此，我们首次尝试对另外两种细胞色素P450酶（CYP109C1和CYP109C2）进行表征，并将它们的生化、生物物理和功能特性与脂肪酸羟化酶CYP109D1的特性进行比较。鉴于脂肪酸的生理重要性，我们研究了CYP109家族所有成员对饱和脂肪酸的结合和转化情况。使用生物传感器测量比较了CYP109家族成员与不同的自体/异体氧化还原伙伴之间的相互作用，结果发现只有CYP109D1与牛肾上腺皮质铁氧化还原蛋白（Adx）形成了复合物。令人惊讶的是，这种相互作用与Adx与其哺乳动物氧化还原伙伴之间的相互作用效率相似。体外重组试验表明，使用CYP109C1时没有活性，尽管已证明其能结合底物；同样，使用CYP109C2和CYP109D1时，饱和脂肪酸会发生亚末端羟基化，其中CYP109D1是一种效率更高的脂肪酸羟化酶。有趣的是，使用长链脂肪酸时，羟基化位置会在脂肪酸链内部移动，从而产生具有生理重要性的产物的可能前体。
