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Structural studies of the regulatory domain of bovine soluble guanylate cyclase
Author(s) -
Garcin Elsa D,
Rassouli-Taylor Leida
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.573.6
Subject(s) - gucy1a3 , gucy1b3 , nitric oxide , mutagenesis , chemistry , heme , gtp' , guanylate cyclase , guanosine triphosphate , biochemistry , gucy2d , protein subunit , soluble guanylyl cyclase , guanylate cyclase 2c , biophysics , microbiology and biotechnology , receptor , enzyme , biology , mutation , organic chemistry , gene
Soluble guanylate cyclase (sGC) converts GTP to cGMP to promote vasodilation. The β subunit contains a heme nitric oxide/oxygen‐binding (HNOX) regulatory domain, an HNOX‐associated (HNOXA) and coiled‐coil dimerization domain, and a catalytic guanylate cyclase domain. Nitric oxide (NO) binds to the heme and allosterically stimulates the cyclase activity. We aim to determine the mechanism of NO‐induced activation of sGC. Ultimately, our results will facilitate the development of novel sGC activators to treat cardiovascular diseases. We focused on two truncated constructs encompassing either the HNOX domain or both HNOX‐HNOXA domains. We used site‐directed mutagenesis to generate proteins that would be highly expressed and soluble in E. coli . From the 18 constructs generated, only 5 were expressed and soluble. Our preliminary results suggest that the HNOXA domain plays a role in proper folding of the HNOX domain and/or stability of the heme, in contrast to what was shown for bacterial HNOX homologs. Crystals of HNOX‐sGC diffracted poorly at the synchrotron, but to our knowledge, these are the first mammalian HNOX crystals. We are now working to improve the diffraction quality of these crystals. Funding: American Heart Association Scientist Development Grant and UMBC DRIF.