Refined structure of cytochrome b562 from Escherichia coli at 1.4 A resolution

Abstract

The structure of cytochrome b562 from Escherichia coli has been refined at 1.4 A resolution against X-ray data collected on a Picker four-circle diffractometer. The triclinic unit cell parameters are a = 33.68 A, b = 50.48 A, c = 32.67 A, alpha = 102.51 degrees, beta = 86.56 degrees and gamma = 107.01 degrees and there are two molecules in the asymmetric unit. A total of 138 cycles of restrained crystallographic refinement using the program PROLSQ were augmented at intermediate stages by two cycles of simulated annealing refinement using X-PLOR. The final crystallographic R-factor is 16.4% for data in the resolution range 6.0 A to 1.4 A for a model containing 1650 protein atoms, 86 heme atoms, 165 water molecules and four sulfate anions. The root-mean-square deviations from ideal bond lengths and angles are 0.012 A and 2.0 degrees, respectively. Each molecule consists of a bundle of four alpha-helices arranged in a simple up-down-up-down manner with a non-covalently bound heme group inserted between the first and fourth helices. In addition, there is a very short 3(10) helix in the 15-residue loop connecting the first and second pairs of helices. The two independent molecules show r.m.s. differences of 0.30 A for main-chain atoms and 0.88 A for all atoms. A detailed comparison with the structurally similar cytochrome c' from Rhodospirulum molishianum is presented. In addition, the titration behavior of cytochrome b562 in solution is discussed in terms of its molecular structure.