Unfolding of apomyoglobin examined by synchrotron footprinting.
A new method to examine the structure and stability of proteins using footprinting is applied to examine the unfolding of apomyoglobin. Unlike previous cleavage based footprinting methods, synchrotron X-ray protein footprinting is based on a quantitative determination of the extent and the site of radiolytic modification of amino acid side chains, analyzed using mass spectrometry. The amino acids most susceptible to radiolytic oxidation (cysteine, methionine, phenylalanine, tyrosine, tryptophan, histidine, proline, and leucine) serve as convenient probes of protein structure to monitor changes in solvent accessibility. To determine if the technique can measure quantitative properties of proteins relevant to structure and function, we examined the equilibrium unfolding of apomyoglobin in urea and compared the results to data derived from fluorescence studies under the same conditions.
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Center for Synchrotron Biosciences, Department of Physiology, Albert Einstein College of Medicine, Yeshiva University, 1300 Morris Park Avenue, Bronx, New York 10461, USA. mrc@aecom.yu.edu
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D001059:Apoproteins / Q000737:chemistry* / Q000378:metabolism; D001671:Biochemistry / Q000379:methods*; D055592:Biophysical Phenomena; D001703:Biophysics; D006418:Heme / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009211:Myoglobin / Q000737:chemistry* / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011489:Protein Denaturation; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D013997:Time Factors; D014508:Urea / Q000494:pharmacology; D014965:X-Rays
D000596:Amino Acids; D001059:Apoproteins; D009211:Myoglobin; C010857:apomyoglobin; D006418:Heme; D014508:Urea; D010100:Oxygen