Investigators at Laval University publish new data on biochemistry
2008 JAN 14 -- "The 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," researchers in Canada report. "A previous study reported that changing Y(CD1) for a Phe causes a significant increase in the 02 combination rate, but almost no change in the O-2 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 O-2 and CO, as evidenced by the increase of the Fe - O-2 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 O-2 and CO. Also, the rate of O-2 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]," wrote H. Ouellet and colleagues, Laval University. The researchers concluded: "Taken together, these data indicate a pivotal role for the W(G8) residue in O-2 binding and stabilization." Ouellet and colleagues published their study in Biochemistry (The roles of Tyr(CD1) and Trp(G8) in Mycobacterium tuberculosis truncated hemoglobin O in ligand binding and on the heme distal site architectures. Biochemistry, 2007;46(41):11440-11450). For additional information, contact M. Guertin, Laval University, Dept. of Biochemistry & Microbiology, Pavillon Marchand, Room 3145, Ste. Foy, PQ G1K 7P4, Canada. Publisher contact information for the journal Biochemistry is: American Chemical Society, 1155 16th St., NW, Washington, DC 20036, USA. Keywords: Canada, Biochemistry, Cutaneous Tuberculosis, Mutagenesis, Mycobacteria, Mycobacterium Tuberculosis, Spectroscopy, Surgery, Laval University. This article was prepared by Tuberculosis Week editors from staff and other reports. Copyright 2008, Tuberculosis Week via NewsRx.com.
|
