1b5m
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(New page: 200px<br /><applet load="1b5m" size="450" color="white" frame="true" align="right" spinBox="true" caption="1b5m, resolution 2.7Å" /> '''RAT OUTER MITOCHONDRI...)
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Revision as of 09:15, 20 November 2007
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RAT OUTER MITOCHONDRIAL MEMBRANE CYTOCHROME B5
Overview
The role played by the outer mitochondrial membrane (OM) cytochrome b5, heme propionate groups in the electrostatic binding between OM cytochrome, b5 and horse heart cytochrome c was investigated by 13C NMR spectroscopy, and X-ray crystallography. To achieve these aims, 13C-labeled heme OM, cytochrome b5 was expressed in Escherichia coli as previously described, [Rivera M., Walker, F.A. (1995) Anal. Biochem. 230, 295-302]. Assignment, of the resonances arising from the heme propionate carbons in, ferricytochrome b5 was carried out by a combination of one- and, two-dimensional NMR experiments. Titrations of [13C]heme-labeled OM, cytochrome b5 with horse heart cytochrome c were carried out in order to, monitor the resonances arising from the heme propionate carbonyl carbons, in OM cytochrome b5. The results from these titrations clearly show that, only the heme propionate located on the exposed heme edge in OM cytochrome, b5 participates in the electrostatic stabilization of the complex between, OM cytochrome b5 and horse heart cytochrome c. Similar experiments carried, out monitoring 13C resonances arising from several other heme substituents, demonstrated that the stoichiometry of the complex is 1:1. A conditional, binding constant, K which equals 3.8 x 10(4) +/- 1.4 x 10(4) at mu = 0.02, M, was obtained for the formation of the complex by fitting the binding, curves obtained experimentally to a model based on this stoichiometry. The, X-ray crystal structure of rat liver OM cytochrome b5 solved to 2.7 A, resolution shows that the structures of bovine liver microsomal cytochrome, b5 and rat liver OM cytochrome b5 are almost identical when compared at, medium resolution. The similarity between the two structures, combined, with the findings that only the heme propionate located on the exposed, heme edge of OM cytochrome b5 participates in the electrostatic binding to, cytochrome c and that the stability of this complex is similar to that, measured for the association between microsomal cytochrome b5 and, cytochrome c, clearly indicates that the site of interaction on OM, cytochrome b5 is almost identical to the one elucidated for microsomal, cytochrome b5. It is therefore possible to conclude that the large body of, information gathered by many investigators for the nonphysiological, interaction between microsomal cytochrome b5 and cytochrome c (recently, reviewed) [Mauk, A. G. Mauk, M. R., Moore, G. R., & Northrup, S. H. (1995), Bioenerg. Biomembr. 27, 311-330] has indeed biological as well as, pedagogical validity.
About this Structure
1B5M is a Single protein structure of sequence from Rattus norvegicus with HEM as ligand. Full crystallographic information is available from OCA.
Reference
13C NMR spectroscopic and X-ray crystallographic study of the role played by mitochondrial cytochrome b5 heme propionates in the electrostatic binding to cytochrome c., Rodriguez-Maranon MJ, Qiu F, Stark RE, White SP, Zhang X, Foundling SI, Rodriguez V, Schilling CL 3rd, Bunce RA, Rivera M, Biochemistry. 1996 Dec 17;35(50):16378-90. PMID:8973214
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