2qrw
Crystal structure of Mycobacterium tuberculosis trHbO WG8F mutant
Structural highlights
FunctionTRHBO_MYCTU When expressed in E.coli and M.smegmatis, HbO increases oxygen uptake. Membrane vesicles of E.coli carrying HbO show a respiration activity about twice that of membranes without HbO. HbO seems to interact with a terminal oxidase. Therefore, HbO could participate in oxygen/electron-transfer process, suggesting a function related to the facilitation of oxygen transfer during aerobic metabolism of M.tuberculosis. Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedThe crystal structure of the cyano-met form of Mt-trHbO revealed two unusual distal residues Y(CD1) and W(G8) forming a hydrogen-bond network with the heme-bound ligand [Milani, M., et al. (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 5766-5771]. W(G8) is an invariant residue in group II and group III trHbs and has no counterpart in other globins. A previous study reported that changing Y(CD1) for a Phe causes a significant increase in the O2 combination rate, but almost no change in the O2 dissociation rate [Ouellet, H., et al. (2003) Biochemistry 42, 5764-5774]. Here we investigated the role of the W(G8) in ligand binding by using resonance Raman spectroscopy, stopped-flow spectrophotometry, and X-ray crystallography. For this purpose, W(G8) was changed, by site-directed mutagenesis, to a Phe in both the wild-type protein and the mutant Y(CD1)F to create the single mutant W(G8)F and the double mutant Y(CD1)F/W(G8)F, respectively. Resonance Raman results suggest that W(G8) interacts with the heme-bound O2 and CO, as evidenced by the increase of the Fe-O2 stretching mode from 559 to 564 cm-1 and by the lower frequency of the Fe-CO stretching modes (514 and 497 cm-1) compared to that of the wild-type protein. Mutation of W(G8) to Phe indicates that this residue controls ligand binding, as evidenced by a dramatic increase of the combination rates of both O2 and CO. Also, the rate of O2 dissociation showed a 90-1000-fold increase in the W(G8)F and Y(CD1)F/W(G8)F mutants, that is in sharp contrast with the values obtained for the other distal mutants Y(B10)F and Y(CD1)F [Ouellet, H., et al. (2003) Biochemistry 42, 5764-5774]. Taken together, these data indicate a pivotal role for the W(G8) residue in O2 binding and stabilization. The roles of Tyr(CD1) and Trp(G8) in Mycobacterium tuberculosis truncated hemoglobin O in ligand binding and on the heme distal site architecture.,Ouellet H, Milani M, LaBarre M, Bolognesi M, Couture M, Guertin M Biochemistry. 2007 Oct 16;46(41):11440-50. Epub 2007 Sep 22. PMID:17887774[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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