The crystal structure of peroxymyoglobin generated through cryoradiolytic reduction of myoglobin compound III during data collection.
Myoglobin has the ability to react with hydrogen peroxide, generating high-valent complexes similar to peroxidases (compounds I and II), and in the presence of excess hydrogen peroxide a third intermediate, compound III, with an oxymyoglobin-type structure is generated from compound II. The compound III is, however, easily one-electron reduced to peroxymyoglobin by synchrotron radiation during crystallographic data collection. We have generated and solved the 1.30 A (1 A=0.1 nm) resolution crystal structure of the peroxymyoglobin intermediate, which is isoelectric to compound 0 and has a Fe-O distance of 1.8 A and O-O bond of 1.3 A in accordance with a Fe(II)-O-O- (or Fe(III)-O-O2-) structure. The generation of the peroxy intermediate through reduction of compound III by X-rays shows the importance of using single-crystal microspectrophotometry when doing crystallography on metalloproteins. After having collected crystallographic data on a peroxy-generated myoglobin crystal, we were able (by a short annealing) to break the O-O bond leading to formation of compound II. These results indicate that the cryoradiolytic-generated peroxymyoglobin is biologically relevant through its conversion into compound II upon heating. Additionally, we have observed that the Xe1 site is occupied by a water molecule, which might be the leaving group in the compound II to compound III reaction.
Hersleth
Hans-Petter
HP
University of Oslo, Department of Molecular Biosciences, P.O. Box 1041 Blindern, N-0316 Oslo, Norway.
Hsiao
Ya-Wen
YW
Ryde
Ulf
U
Görbitz
Carl Henrik
CH
Andersson
K Kristoffer
KK
D000818:Animals; D018360:Crystallography, X-Ray; D006736:Horses; D006861:Hydrogen Peroxide / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009211:Myoglobin / Q000737:chemistry* / Q000378:metabolism; D016877:Oxidants / Q000737:chemistry; D011487:Protein Conformation; D014867:Water / Q000737:chemistry
D009211:Myoglobin; D016877:Oxidants; C048685:peroxymyoglobin; D014867:Water; D006861:Hydrogen Peroxide