Activation of soluble guanylate cyclase by NO-hemoproteins involves NO-heme exchange. Comparison of heme-containing and heme-deficient enzyme forms.
Activation of soluble guanylate cyclase by NO-hemoproteins involves NO-heme exchange. Comparison of heme-containing and heme-deficient enzyme forms.
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发表时间：
1986-04
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通讯作者：
L. Ignarro;J. Adams;P. M. Horwitz;K. S. Wood
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作者：
L. Ignarro;J. Adams;P. M. Horwitz;K. S. Wood
本文报道了高分子量亚硝基血红素蛋白复合物对牛肺中纯化的可溶性鸟苷酸环化酶的激活机理。研究了含血红素、血红素缺乏和血红素重建形式的鸟苷酸环化酶。一氧化氮（NO）和亚硝基化合物激活含血红素和血红素重组酶（超过50倍），伴随着Soret吸收峰从431至398 nm的位移，但未能激活或改变血红素缺乏酶的光谱特性。相反，预先形成的NO-血红素蛋白复合物以及低分子量NO-血红素激活所有形式的鸟苷酸环化酶。血红素缺乏鸟苷酸环化酶首先与过量的NO-血红蛋白、NO-肌红蛋白或NO-过氧化氢酶反应，然后通过柱色谱法从NO-血红素蛋白中快速分离。分光光度分析表明，NO-血红素部分被转移从每个NO-血红素蛋白血红素缺乏鸟苷酸环化酶。每摩尔全酶结合约1摩尔NO-血红素，这种酶形式的比活性比未反应的血红素缺陷酶高50倍以上。NO-血红素与鸟苷酸环化酶紧密结合，因为酶结合的NO-血红素没有转移到脱辅基血红蛋白中。NO-血红素蛋白激活的酶在动力学上与NO或NO-血红素激活的酶非常相似。相反，血红素缺乏鸟苷酸环化酶和血红素蛋白之间的反应并不导致血红素转移，而血红素单独迅速重建的酶。这些观察结果表明，可溶性鸟苷酸环化酶可以很容易地重建，从而激活，NO-血红素通过交换反应与NO-血红素蛋白。
The mechanism of activation of soluble guanylate cyclase purified from bovine lung by high molecular weight, nitrosyl-hemoprotein complexes is reported. Heme-containing, heme-deficient, and heme-reconstituted forms of guanylate cyclase were studied. Nitric oxide (NO) and nitroso compounds activated heme-containing and heme-reconstituted enzymes (over 50-fold), with an accompanying shift in the Soret absorption peak from 431 to 398 nm, but failed to activate or alter the spectral characteristics of heme-deficient enzyme. In contrast, preformed NO-hemoprotein complexes as well as low molecular weight NO-heme activated all forms of guanylate cyclase. Heme-deficient guanylate cyclase was first reacted with excess amounts of NO-hemoglobin, NO-myoglobin, or NO-catalase and then rapidly separated from the NO-hemoprotein by column chromatography. Spectrophotometric analysis indicated that the NO-heme moiety was transferred from each of the NO-hemoproteins to heme-deficient guanylate cyclase. Approximately 1 mol of NO-heme was bound per mol of holoenzyme and the specific activity of this enzyme form was over 50-fold greater than that of unreacted, heme-deficient enzyme. NO-heme was tightly bound to guanylate cyclase as no transfer of enzyme-bound NO-heme to apohemoglobin was evident. Enzyme activated by NO-hemoproteins closely resembled, kinetically, that activated by NO or NO-heme. In contrast, reactions between heme-deficient guanylate cyclase and hemoproteins did not result in heme transfer, whereas heme alone rapidly reconstituted the enzyme. These observations indicate that soluble guanylate cyclase can be readily reconstituted with, and thereby activated by, NO-heme through an exchange reaction with NO-hemoproteins.