1m20
|
Crystal Structure of F35Y Mutant of Trypsin-solubilized Fragment of Cytochrome b5
OverviewOverview
Conserved phenylalanine 35 is one of the hydrophobic patch residues on the, surface of cytochrome b5 (cyt b5). This patch is partially exposed on the, surface of cyt b5 while its buried face is in direct van der Waals', contact with heme b. Residues Phe35 and Phe/Tyr74 also form an aromatic, channel with His39, which is one of the axial ligands of heme b. By, site-directed mutagenesis we have produced three mutants of cyt b5:, Phe35-->Tyr, Phe35-->Leu, and Phe35-->His. We found that of these three, mutants, the Phe35-->Tyr mutant displays abnormal properties. The redox, potential of the Phe35-->Tyr mutant is 66 mV more negative than that of, the wild-type cyt b5 and the oxidized Phe35-->Tyr mutant is more stable, towards thermal and chemical denaturation than wild-type cyt b5. In this, study we studied the most interesting mutant, Phe35-->Tyr, by X-ray, crystallography, thermal denaturation, CD and kinetic studies of heme, dissociation to explore the origin of its unusual behaviors. Analysis of, crystal structure of the Phe35-->Tyr mutant shows that the overall, structure of the mutant is basically the same as that of the wild-type, protein. However, the introduction of a hydroxyl group in the heme pocket, and the increased van der Waals' and electrostatic interactions between, the side chain of Tyr35 and the heme probably result in enhancement of, stability of the Phe35-->Tyr mutant. The kinetic difference of the heme, trapped by the heme pocket also supports this conclusion. The detailed, conformational changes of the proteins in response to heat have been, studied by CD for the first time, revealing the existence of the folding, intermediate.
About this StructureAbout this Structure
1M20 is a Single protein structure of sequence from Bos taurus with HEM as ligand. Full crystallographic information is available from OCA.
ReferenceReference
X-ray crystallography, CD and kinetic studies revealed the essence of the abnormal behaviors of the cytochrome b5 Phe35-->Tyr mutant., Yao P, Wu J, Wang YH, Sun BY, Xia ZX, Huang ZX, Eur J Biochem. 2002 Sep;269(17):4287-96. PMID:12199707
Page seeded by OCA on Tue Nov 20 21:04:41 2007