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.
Department of Biological Chemistry, University of Michigan Medical School, 1150 West Medical Center Drive, Ann Arbor, MI 48109-0606, USA.
David P
DP
Ballou
Yunde
Y
Zhao
Philip E
PE
Brandish
Michael A
MA
Marletta
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