THERMUS-THERMOPHILUS CYTOCHROME-C(552) - A NEW HIGHLY THERMOSTABLE CYTOCHROME-C STRUCTURE OBTAINED BY MAD PHASING

The three-dimensional structure of cytochrome-c(552) from Thermus ther mophilus has been determined by the multiple anomalous dispersion tech nique using synchrotron radiation and refined to a resolution of 1.28 Angstrom. Data collection at 90 K and the recording of three data sets (f'-minimum: 7125 eV, f ''-maximum: 7138 eV and reference for scaling : 10,077 eV) resulted in an initial electron density of very high qual ity at 2.1 Angstrom, which was readily interpretable for model buildin g, The model was refined to an R value of 19.1% (R-free = 22.4%) at 1. 28 Angstrom resolution using a fourth data set collected at a photon e nergy of 11,810 eV. Comparison of this thermophilic cytochrome with it s mesophilic mitochondrial or bacterial counterparts reveals significa nt structural differences which are discussed with respect to their im portance for thermostability and binding between this cytochrome and i ts corresponding ba(3)-oxidase. Amino acid sequence similarities to ot her class I cytochromes are very weak and entirely limited to the regi on around the CXXCH motif close to the N terminus. The N-terminal two- thirds of cytochrome-c(552) cover spatial regions around the heme pros thetic group that are similar to those observed for other cytochromes. The actual secondary structural elements that are responsible for tha t shielding do not, however, correlate well to other structures. Only the N-terminal helix (containing the heme binding cysteine residues) a ligns reasonably well with other class I cytochromes. The most strikin g differences that distinguish the present structure from all other cl ass I cytochromes is the C-terminal one-third of the molecule that wra ps around the remainder of the structure as a stabilizing clamp, the e xistence of an extended beta-sheet covering one edge of the heme and t he lack of any internal water molecule. (C) 1997 Academic Press Limite d.