Discovery of stimulator binding to a conserved pocket in the heme domain of soluble guanylyl cyclase
AuthorWales, Jessica A.
Bernier, Sylvie G.
Sheppeck, James E.
Renhowe, Paul A.
Montfort, William R.
AffiliationUniv Arizona, Dept Chem & Biochem
Keywordsguanylate cyclase (guanylyl cyclase)
mass spectrometry (MS)
nuclear magnetic resonance (NMR)
MetadataShow full item record
CitationDiscovery of stimulator binding to a conserved pocket in the heme domain of soluble guanylyl cyclase 2018, 293 (5):1850 Journal of Biological Chemistry
JournalJournal of Biological Chemistry
Rights© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.
Collection InformationThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at email@example.com.
AbstractSoluble guanylyl cyclase (sGC) is the receptor for nitric oxide and a highly sought-after therapeutic target for the management of cardiovascular diseases. New compounds that stimulate sGC show clinical promise, but where these stimulator compounds bind and how they function remains unknown. Here, using a photolyzable diazirine derivative of a novel stimulator compound, IWP-051, and MS analysis, we localized drug binding to the 1 heme domain of sGC proteins from the hawkmoth Manduca sexta and from human. Covalent attachments to the stimulator were also identified in bacterial homologs of the sGC heme domain, referred to as H-NOX domains, including those from Nostoc sp. PCC 7120, Shewanella oneidensis, Shewanella woodyi, and Clostridium botulinum, indicating that the binding site is highly conserved. The identification of photoaffinity-labeled peptides was aided by a signature MS fragmentation pattern of general applicability for unequivocal identification of covalently attached compounds. Using NMR, we also examined stimulator binding to sGC from M. sexta and bacterial H-NOX homologs. These data indicated that stimulators bind to a conserved cleft between two subdomains in the sGC heme domain. L12W/T48W substitutions within the binding pocket resulted in a 9-fold decrease in drug response, suggesting that the bulkier tryptophan residues directly block stimulator binding. The localization of stimulator binding to the sGC heme domain reported here resolves the longstanding question of where stimulators bind and provides a path forward for drug discovery.
Note12 month embargo; published online: 8 December 2017
VersionFinal published version
SponsorsNational Institutes of Health from NIEHS [ES06694]; National Institutes of Health from NCI [CA023074]; BIO5 Institute of the University of Arizona; National Institutes of Health from the National Center for Research Resources (NCRR) [1S10 RR028868-01]; National Institutes of Health from NIGMS [P41GM103399, P41RR002301]; University of Wisconsin-Madison; National Institutes of Health [P41GM103399, S10RR02781, S10RR08438, S10RR023438, S10RR025062, S10RR029220]; National Science Foundation [DMB-8415048, OIA-9977486, BIR-9214394]; U.S. Department of Agriculture