Resolving Complexity in the Interactions of Redox Enzymes and Their Inhibitors: Contrasting Mechanisms for the Inhibition of a Cytochrome c Nitrite Reductase Revealed by Protein Film Voltammetr
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- 作者：James D. Gwyer ; David J. Richardson ; and Julea N. Butt
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：November 30, 2004
- 年：2004
- 卷：43
- 期：47
- 页码：15086 - 15094
- 全文大小：189K
- 年卷期：v.43,no.47(November 30, 2004)
- ISSN：1520-4995
文摘
Cytochrome c nitrite reductase is a dimeric decaheme-containing enzyme that catalyzes thereduction of nitrite to ammonium. The contrasting effects of two inhibitors on the activity of this enzymehave been revealed, and defined, by protein film voltammetry (PFV). Azide inhibition is rapid andreversible. Variation of the catalytic current magnitude describes mixed inhibition in which azide bindsto the Michaelis complex (
bscribe/journals/bichaw/43/i47/eqn/bi049085xe10001.gif"> ~ 40 mM) with a lower affinity than to the enzyme alone (bscribe/journals/bichaw/43/i47/eqn/bi049085xe10002.gif"> ~ 15 mM)and leads to complete inhibition of enzyme activity. The position of the catalytic wave reports tighterbinding of azide when the active site is oxidized (bscribe/journals/bichaw/43/i47/eqn/bi049085xe10003.gif"> ~ 39 
M) than when it is reduced. By contrast,binding and release of cyanide are sluggish. The higher affinity of cyanide for reduced versus oxidizedforms of nitrite reductase is immediately revealed, as is the presence of two sites for cyanide binding andinhibition of the enzyme. Formation of the monocyano complex by reduction of the enzyme followed bya "rapid" scan to high potentials captures the activity-potential profile of this enzyme form and showsit to be distinct from that of the uninhibited enzyme. The biscyano complex is inactive. These studiesdemonstrate the complexity that can be associated with inhibitor binding to redox enzymes and illustratehow PFV readily captures and deconvolves this complexity through its impact on the catalytic propertiesof the enzyme.
M) than when it is reduced. By contrast,binding and release of cyanide are sluggish. The higher affinity of cyanide for reduced versus oxidizedforms of nitrite reductase is immediately revealed, as is the presence of two sites for cyanide binding andinhibition of the enzyme. Formation of the monocyano complex by reduction of the enzyme followed bya "rapid" scan to high potentials captures the activity-potential profile of this enzyme form and showsit to be distinct from that of the uninhibited enzyme. The biscyano complex is inactive. These studiesdemonstrate the complexity that can be associated with inhibitor binding to redox enzymes and illustratehow PFV readily captures and deconvolves this complexity through its impact on the catalytic propertiesof the enzyme.