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Oral presentation

Most structural changes that accompany the binding of NO or YC-1 to soluble guanylyl cyclase (sGC) remain unknown. Because guanylyl cyclases (GCs) and adenylyl cyclases (ACs) are related enzymes, we hypothesized that some mechanisms of regulation of ACs are conserved in GCs.

We conducted a mutational analysis to determine whether:

1. Activation by NO and YC-1 propagate to the catalytic site through interactions between the two subunits of the sGC, as occurs in ACs with forskolin (FSK) and Gsα.

2. YC-1 interacts at a putative pseudo-symmetric pocket that is the analog to the FSK binding pocket of ACs.

Biochemical analysis identified similar phenotypes between analogous AC and GC mutants. Concentration-response curves to NO, YC-1 and both activators reveal that individual activation by YC-1 or NO can be dissociated from synergistic activation. We discovered that YC-1 could synergistically increase NO-stimulated activity without increasing affinity for NO. Thus, YC-1 appears to have 3 modes of action: individual activation, increased NO occupancy and increased NO efficacy.

To account for these findings, a structural model of YC-1 interaction with the sGC is proposed, in which there is an equilibrium between two orientations of bound YC-1 that determine the mode of activation.

Acknowledgements

This work was supported by American Heart Association Grant SDG 0130506T and the Foundation of UMDNJ (A.B)

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