DETERMINATION OF THE METHIONINE HEME LIGAND CONFORMATION IN CYTOCHROME C(H) FROM METHYLOPHILUS-METHYLOTROPHUS BY 2-DIMENSIONAL H-1-NMR

A complete relaxation matrix approach was used to determine the confor mation of the Met ligand to the haem in Methylophilus methylotrophus c ytochrome c(H), including the configuration at the sulphur; the patter n of NOEs is shown to be dominated by spin diffusion at mixing times a s short as 25 ms, rendering the two-spin approximation unreliable. The Met conformation, together with an analysis of aromatic proton resona nces, indicates a structure almost identical with that found by x-ray crystallography for Pseudomonas aeruginosa cytochrome c(551), which ha s a 45% similarity in sequence. The paramagnetic shifts of the haem me thyl proton resonances in the oxidized protein support the view that t he asymmetry in the electronic structure of the haem is dominated by t he orientation of the Met ligand.