The 1H NMR spectrum of the the cyanide adduct of a triply mutated Saccharomyces cerevisiae iso-1-cytochrome c (His39Gln/Met80Ala/Cys102Ser) in the oxidized form has been assigned through 1D NOE and 2D COSY, TOCSY, NOESY, and NOE-NOESY experiments; 562 protons out of a total of 683 have been assigned. The solution structure, the first of a paramagnetic heme protein, was determined using 1426 meaningful NOE constraints out of a total of 1842 measured NOEs. The RMSD values at the stage of restrained energy minimization of 17 structures obtained from distance geometry calculations are 0.68 +/- 0.11 and 1.32 +/- 0.14 A for the backbone and all heavy atoms, respectively. The quality, in terms of RMSD, of the present structure is the same as that obtained for the solution structure of the diamagnetic horse heart ferrocytochrome c [Qi, P. X., et al. (1994) Biochemistry 33, 6408-6419]. The secondary structure elements and the overall folding in the variant are observed to be the same as those of the wild-type protein for which the X-ray structure is available. However, the replacement of the methionine axial ligand with an alanine residue creates a ligand-binding "distal cavity". The properties of the distal cavity seen in this solution structure are compared to those of other heme proteins.

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Journal Article | Research Support, Non-U.S. Gov't

Authors

Banci L, Bertini I, Bren KL, Gray HB, Sompornpisut P, Turano P

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