Cryoradiolytic reduction of crystalline heme proteins: analysis by UV-Vis spectroscopy and X-ray crystallography.
The X-ray crystallographic analysis of redox-active systems may be complicated by photoreduction. Although radiolytic reduction by the probing X-ray beam may be exploited to generate otherwise short-lived reaction intermediates of metalloproteins, it is generally an undesired feature. Here, the X-ray-induced reduction of the three heme proteins myoglobin, cytochrome P450cam and chloroperoxidase has been followed by on-line UV-Vis absorption spectroscopy. All three systems showed a very rapid reduction of the heme iron. In chloroperoxidase the change of the ionization state from ferric to ferrous heme is associated with a movement of the heme-coordinating water molecule. The influence of the energy of the incident X-ray photons and of the presence of scavengers on the apparent reduction rate of ferric myoglobin crystals was analyzed.
Beitlich
Thorsten
T
Max Planck Institute for Medical Research, Department of Biomolecular Mechanisms, Jahnstrasse 29, 69120 Heidelberg, Germany.
Kühnel
Karin
K
Schulze-Briese
Clemens
C
Shoeman
Robert L
RL
Schlichting
Ilme
I
D003460:Crystallization / Q000379:methods*; D018360:Crystallography, X-Ray; D004307:Dose-Response Relationship, Radiation; D005615:Freezing; D006420:Hemeproteins / Q000737:chemistry* / Q000528:radiation effects* / Q000648:ultrastructure; D010084:Oxidation-Reduction / Q000528:radiation effects; D011487:Protein Conformation / Q000528:radiation effects; D011829:Radiation Dosage; D013056:Spectrophotometry, Ultraviolet; D014466:Ultraviolet Rays; D014965:X-Rays
D006420:Hemeproteins