Revisiting the kinetics of nitric oxide (NO) binding to soluble guanylate cyclase: the simple NO-binding model is incorrect.
Soluble guanylate cyclase (sGC) is a ferrous iron hemoprotein receptor for nitric oxide (NO). NO binding to the heme activates the enzyme 300-fold. sGC as isolated is five-coordinate, ferrous with histidine as the axial ligand. The NO-activated enzyme is a five-coordinate nitrosyl complex where the axial histidine bond is broken. Past studies using rapid-reaction kinetics demonstrated that both the formation of a six-coordinate intermediate and the conversion of the intermediate to the activated five-coordinate nitrosyl complex depended on the concentration of NO. A model invoking a second NO molecule as a catalyst for the conversion of the six-coordinate intermediate to the five-coordinate sGC-NO complex was proposed to explain the observed kinetic data. A recent study [Bellamy, T. C., Wood, J. & Garthwaite, J. (2002) Proc. Natl. Acad. Sci. USA 99, 507-510] concluded that a simple two-step binding model explains the results. Here we show through further analysis and simulations of previous data that the simple two-step binding model cannot be used to describe our results. Instead we show that a slightly more complex two-step binding model, where NO is used as a ligand in the first step and a catalyst in the second step, can describe our results quite satisfactorily. These new simulations combined with the previous activation data lead to the conclusion that the intermediate six-coordinate sGC-NO complex has substantial activity. The model derived from our simulations also can account for the slow deactivation of sGC that has been observed in vitro.
Ballou
David P
DP
Department of Biological Chemistry, University of Michigan Medical School, 1150 West Medical Center Drive, Ann Arbor, MI 48109-0606, USA.
Zhao
Yunde
Y
Brandish
Philip E
PE
Marletta
Michael A
MA
D000421:Alcaligenes / Q000378:metabolism; D003198:Computer Simulation; D003574:Cytochrome c Group / Q000378:metabolism; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008954:Models, Biological; D009569:Nitric Oxide / Q000378:metabolism*; D015398:Signal Transduction; D012995:Solubility; D013053:Spectrophotometry
D003574:Cytochrome c Group; D009569:Nitric Oxide; D006162:Guanylate Cyclase