Berka V, Palmer G, Chen P F, Tsai A L
Division of Hematology, Department of Internal Medicine, University of Texas Medical School at Houston 77030, USA.
Biochemistry. 1998 Apr 28;37(17):6136-44. doi: 10.1021/bi980133l.
We have evaluated the influence of a series of substituted imidazoles on the heme structure of endothelial nitric oxide synthase (eNOS). Optical, MCD, and EPR spectra reveal widely differing effects on heme spin state and geometry. 1-Substituted imidazoles always yield low-spin heme complexes, but the size of the 2- and 4-substituent influences their structural effects on the heme. Methyl substituents lead to low-spin complexes while the bulky phenyl group yields high-spin complexes. The only exception to this behavior is provided by 2-aminoimidazole. Although this compound has three functional groups which can serve as an axial ligand to the heme, its binding to eNOS leads to a pure high-spin complex. This result can only be interpreted as due to a direct binding of 2-aminoimidazole to the guanidine binding subdomain of L-arginine. MCD spectra also imply that an O-ligand is present in the low-spin resting eNOS, while EPR data reveal the presence of two low-spin heme complexes in resting eNOS and its imidazole complexes. EPR also distinguishes four different high-spin forms of eNOS generated by different imidazole analogues. This series of ligands promises to be useful in probing the subtle structural difference among the active sites of three NOS isozymes and in developing selective inhibitors to these important enzymes.
我们评估了一系列取代咪唑对内皮型一氧化氮合酶（eNOS）血红素结构的影响。光学、磁圆二色性（MCD）和电子顺磁共振（EPR）光谱揭示了对血红素自旋状态和几何结构的广泛不同影响。1-取代咪唑总是产生低自旋血红素配合物，但2-和4-取代基的大小影响它们对血红素的结构效应。甲基取代基导致低自旋配合物，而庞大的苯基产生高自旋配合物。这种行为的唯一例外是2-氨基咪唑。尽管该化合物有三个可作为血红素轴向配体的官能团，但其与eNOS的结合导致纯高自旋配合物。该结果只能解释为2-氨基咪唑直接与L-精氨酸的胍结合亚结构域结合。MCD光谱还表明在低自旋静止eNOS中存在一个O-配体，而EPR数据揭示在静止eNOS及其咪唑配合物中存在两种低自旋血红素配合物。EPR还区分了由不同咪唑类似物产生的四种不同的高自旋形式的eNOS。这一系列配体有望用于探究三种一氧化氮合酶同工酶活性位点之间的细微结构差异，并用于开发针对这些重要酶的选择性抑制剂。
