8rkp

Publication Abstract from PubMed

Cytochromes c'-alpha are nitric oxide (NO)-binding heme proteins derived from bacteria that can thrive in a wide range of temperature environments. Studies of mesophilic Alcaligenes xylosoxidans cytochrome c'-alpha (AxCP-alpha) have revealed an unusual NO-binding mechanism involving both heme faces, in which NO first binds to form a distal hexa-coordinate Fe(II)-NO (6cNO) intermediate and then displaces the proximal His to form a proximal penta-coordinate Fe(II)-NO (5cNO) final product. Here, we characterize a thermally stable cytochrome c'-alpha from thermophilic Hydrogenophilus thermoluteolus (PhCP-alpha) to understand how protein thermal stability affects NO binding. Electron paramagnetic and resonance Raman spectroscopies reveal the formation of a PhCP-alpha 5cNO product, with time-resolved (stopped-flow) UV-vis absorbance indicating the involvement of a 6cNO intermediate. Relative to AxCP-alpha, the rates of 6cNO and 5cNO formation in PhCP-alpha are approximately 11- and approximately 13-fold lower, respectively. Notably, x-ray crystal structures of PhCP-alpha in the presence and absence of NO suggest that the sluggish formation of the proximal 5cNO product results from conformational rigidity: the Arg-132 residue (adjacent to the proximal His ligand) is held in place by a salt bridge between Arg-75 and Glu-135 (an interaction not present in AxCP-alpha or a psychrophilic counterpart). Overall, our data provide fresh insights into structural factors controlling NO binding in heme proteins, including 5cNO complexes relevant to eukaryotic NO sensors.

Conformational rigidity of cytochrome c'-alpha from a thermophile is associated with slow NO binding.,Fujii S, Wilson MT, Adams HR, Mikolajek H, Svistunenko DA, Smyth P, Andrew CR, Sambongi Y, Hough MA Biophys J. 2024 Aug 20;123(16):2594-2603. doi: 10.1016/j.bpj.2024.06.026. Epub , 2024 Jun 26. PMID:38937973^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.