Kishore N S, Wood D C, Mehta P P, Wade A C, Lu T, Gokel G W, Gordon J I
Department of Infectious Diseases, G. D. Searle Co., St. Louis, Missouri.
J Biol Chem. 1993 Mar 5;268(7):4889-902.
Human myristoyl-CoA synthetase and myristoyl-CoA:protein N-myristoyltransferase (hNmt) have been partially purified from an erythroleukemia cell line. Their substrate specificities were examined using two in vitro assays of enzyme activity together with a panel of C7-C17 saturated fatty acids plus 72 myristic acid analogs containing oxygen, sulfur, ketocarbonyl, ester, amide, cis and trans double bonds, triple bonds, and para-substituted phenyl groups. There is an inverse relationship between the polarity and the activity of C14 fatty acid substrates of myristoyl-CoA synthetase. Surveys of tetradecenoic and tetradecynoic acids suggest that myristate is bound to the synthetase in a bent conformation with a principal bend occurring in the vicinity of C5-C6. The synthetase can tolerate a somewhat wider range of physical chemical properties in acyl chains than can the monomeric hNmt. However, like myristoyl-CoA synthetase, there is an inverse relationship between acyl chain polarity and the activities of hNmt's acyl-CoA substrates. Moreover, the acyl chain of myristoyl-CoA appears to be bound to hNmt in a bent conformation with bends located in the vicinity of C5 and C8. The acyl chain specificities of both enzymes make them well suited to utilize efficiently any cellular pools of 5Z-tetradecenoic and 5Z,8Z-tetradecadienoic acids and their CoA derivatives. This feature may account for the recent observation that in some mammalian cell lineages, certain N-myristoyl-proteins are heterogeneously acylated with these C14 fatty acids. Finally, the acyl-CoA binding sites of human and Saccharomyces cerevisiae Nmts appear to have been highly conserved. Given their overlapping yet distinct peptide substrate specificities, development of species-specific inhibitors of Nmts should probably focus on structural features recognized in the enzymes' peptide substrates rather than in the acyl chain of their acyl-CoA substrates.
人豆蔻酰辅酶A合成酶和豆蔻酰辅酶A：蛋白质N-豆蔻酰转移酶（hNmt）已从一种红白血病细胞系中部分纯化出来。利用两种体外酶活性测定方法以及一组C7 - C17饱和脂肪酸加上72种含有氧、硫、酮羰基、酯、酰胺、顺式和反式双键、三键以及对位取代苯基的肉豆蔻酸类似物，对它们的底物特异性进行了检测。豆蔻酰辅酶A合成酶的C14脂肪酸底物的极性与其活性之间存在反比关系。对十四碳烯酸和十四碳炔酸的研究表明，肉豆蔻酸以弯曲构象与合成酶结合，主要弯曲发生在C5 - C6附近。与单体hNmt相比，合成酶对酰基链物理化学性质的耐受范围稍宽一些。然而，与豆蔻酰辅酶A合成酶一样，hNmt的酰基辅酶A底物的酰基链极性与其活性之间也存在反比关系。此外，豆蔻酰辅酶A的酰基链似乎以弯曲构象与hNmt结合，弯曲位于C5和C8附近。两种酶的酰基链特异性使它们非常适合有效利用5Z - 十四碳烯酸和5Z,8Z - 十四碳二烯酸及其辅酶A衍生物的任何细胞池。这一特征可能解释了最近的观察结果，即在某些哺乳动物细胞谱系中，某些N - 豆蔻酰化蛋白被这些C14脂肪酸异质酰化。最后，人和酿酒酵母Nmts的酰基辅酶A结合位点似乎高度保守。鉴于它们重叠但又不同的肽底物特异性，开发Nmts的物种特异性抑制剂可能应该侧重于酶的肽底物中识别的结构特征，而不是其酰基辅酶A底物的酰基链。
