Resonance Raman evidence for the presence of two heme pocket conformations with varied activities in CO-bound bovine soluble guanylate cyclase and their conversion
Resonance Raman evidence for the presence of two heme pocket conformations with varied activities in CO-bound bovine soluble guanylate cyclase and their conversion
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DOI：
10.1021/bi0489208
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发表时间：
2005-01-25
期刊：
影响因子：
2.9
通讯作者：
Kitagawa, T
中科院分区：
文献类型：
--
作者：
Li, ZQ;Pal, B;Kitagawa, T
5个独立研究小组报道的可溶性鸟苷环化酶(SGC)的共振拉曼光谱分为两种类型：sGC(1)和sGC(2)。在此，我们证明了从牛肺分离的sGC的RR谱只含有sGC(2)，而这两种物质都是在CO结合形式(CO-sGC)的光谱中观察到的。随着异源化合物YC-1的加入，这两种形式的相对布居数从最初的CO-sGC(2)为主，Fe-CO(nu(Fe-CO))和C-O伸缩模式(nu(CO))分别为472和1985 cm(-1)，转变为以CO-sGC(1)为主，nu(Fe-CO)和nu(CO)分别为488和1969 cm(-1)。进一步添加底物GTP完成了这一改变。国内生产总值和cGMP的作用明显较弱，而底物类似物GTP-伽马-S的作用与GTP相似。而三磷酸腺苷则相反，并能抑制YC-1和GTP的作用。在YC-1和GTP同时存在的情况下，血红素的乙烯基振动受到显著影响。在521、488、363和227 cm(-1)处观察到新的CO同位素敏感带。521 cm(-1)谱带归属于CTAB胶束中铁原卟啉IX模型化合物的五配位(5c)物种。与472 cm(-1)物种不同的是，当使用普通的拉曼自旋池时，488和521 cm(-1)物种都明显不能光解，这表明光解离的CO迅速复合。在这些发现的基础上，提出了YC-1与血红素口袋的结合。
Resonance Raman (RR) spectra of soluble guanylate cyclase (sGC) reported by five independent research groups have been classified as two types: sGC(1) and sGC(2). Here we demonstrate that the RR spectra of sGC isolated from bovine lung contain only sGC(2) while both species are observed in the spectra of the CO-bound form (CO-sGC). The relative populations of the two forms were altered from an initial composition in which the CO-sGC(2) form predominated, with the Fe-CO (nu(Fe-CO)) and C-O stretching modes (nu(CO)) at 472 and 1985 cm(-1), respectively, to a composition dominated by the CO-sGC(1) form with nu(Fe-CO) and nu(CO) at 488 and 1969 cm(-1), respectively, following the addition of a xenobiotic, YC-1. Further addition of a substrate, GTP, completed the change. GDP and cGMP had a significantly weaker effect, while a substrate analogue, GTP-gamma-S, had an effect similar to that of GTP. In contrast, ATP had a reverse effect, and suppressed the effects of YC-1 and GTP. In the presence of both YC-1 and GTP, vinyl vibrations of heme were significantly influenced. New CO isotope-sensitive bands were observed at 521, 488, 363, and 227 cm(-1). The 521 cm(-1) band was assigned to the five-coordinate (5c) species from the model compound studies using ferrous iron protoporphyrin IX in CTAB micelles. Distinct from the 472 cm(-1) species, both the 488 and 521 cm(-1) species were apparently un-photodissociable when an ordinary Raman spinning cell was used, indicating rapid recombination of photodissociated CO. On the basis of these findings, binding of YC-1 to the heme pocket is proposed.