Quantitative structural comparisons of heme protein crystals and solutions using resonance Raman spectroscopy.
Resonance Raman difference spectra have been used to compare crystal and solution samples of metmyoglobin (metMb), deoxymyoglobin (deoxyMb), and cytochrome P450. At pH 6.0, the frequency shifts of the heme core size sensitive bands v2, v3, and v4 were determined to be less than 0.3, 1.0, and 0.3 cm-1, respectively, for metMb and to be less than 1.0, 1.0, and 0.3 cm-1, respectively, for deoxyMb. This shows that the heme core size differences between the crystal and solution conformations are less than 0.002 A for metMb and less than 0.003 A for deoxyMb. These results disagree with a recent extended X-ray absorption fine structure study [Zhang, K., Chance, B., Reddy, K. S., Ayene, I., Stern, E. A., & Bunker, G. (1991) Biochemistry 30, 9116-9120] which claims that a 0.05-A difference exists in the average iron-ligand distance between the crystalline and solution forms of metMb at pH 6.5. At pH 8.5, metMb solution samples change gradually from a predominantly high-spin to a predominantly low-spin species as the ammonium sulfate concentration is increased to the level found in the crystal mother liquor. No Raman frequency shifts are found between the crystal and solution forms of metMb at pH 8.5 when the ammonium sulfate concentrations are equal. On the other hand, for deoxyMb, we find a significant alteration in the 220/240-cm-1 line shape and relative intensities, suggesting that some histidine-heme perturbation takes place upon crystallization.(ABSTRACT TRUNCATED AT 250 WORDS)
Zhu
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Department of Physics, Northeastern University, Boston, Massachusetts 02115.
Sage
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Champion
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D000645:Ammonium Sulfate / Q000494:pharmacology; D003460:Crystallization; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry; D006420:Hemeproteins / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008786:Metmyoglobin / Q000737:chemistry; D009211:Myoglobin / Q000031:analogs & derivatives / Q000737:chemistry; D010782:Photolysis; D012996:Solutions; D013057:Spectrum Analysis; D013059:Spectrum Analysis, Raman; D014965:X-Rays
D006420:Hemeproteins; D009211:Myoglobin; D012996:Solutions; C055547:deoxymyoglobin; D008786:Metmyoglobin; D003577:Cytochrome P-450 Enzyme System; D000645:Ammonium Sulfate