2qrw

From Proteopedia

Revision as of 10:23, 23 January 2008

Crystal stucture of Mycobacterium tuberculosis trHbO WG8F mutant

Overview

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. 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.

About this Structure

2QRW is a Single protein structure of sequence from Mycobacterium tuberculosis with , and as ligands. Full crystallographic information is available from OCA.

Reference

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

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