Mechanism of Binding of NO to Soluble Guanylyl Cyclase: Implication for the Second NO Binding to the Heme Proximal Site

详细信息    查看全文

• 作者：Emil Martin ; Vladimir Berka ; Iraida Sharina ; Ah-Lim Tsai
• 刊名：Biochemistry
• 出版年：2012
• 出版时间：April 3, 2012
• 年：2012
• 卷：51
• 期：13
• 页码：2737-2746
• 全文大小：445K
• 年卷期：v.51,no.13(April 3, 2012)
• ISSN：1520-4995

文摘

Soluble guanylyl cyclase (sGC), the key enzyme for the formation of second messenger cyclic GMP, is an authentic sensor for nitric oxide (NO). Binding of NO to sGC leads to strong activation of the enzyme activity. Multiple molecules and steps of binding of NO to sGC have been implicated, but the target of the second NO and the detailed binding mechanism remain controversial. In this study, we used ¹⁵NO and ¹⁴NO and anaerobic sequential mixing鈥揻reeze-quench electron paramagnetic resonance to unambiguously confirm that the heme Fe is the target of the second NO. The linear dependence on NO concentration up to 600 s^鈥? for the observed rate of the second step of NO binding not only indicates that the binding site of the second NO is different from that in the first step, i.e., the proximal site of the heme, but also supports a concerted mechanism in which the dissociation of the His105 proximal ligand occurs simultaneously with the binding of the second NO molecule. Computer modeling successfully predicts the kinetics of formation of a set of five-coordinate NO complexes with the ligand on either the distal or proximal site and supports the selective release of NO from the distal side of the transient bis-NO鈥搒GC complex. Thus, as has been demonstrated with cytochrome c鈥? a five-coordinate NO鈥搒GC complex containing a proximal NO is formed after the binding of the second NO.