A method for probing the topography and interactions of proteins: footprinting of myoglobin.
We describe a procedure for mapping residues on the surface of a protein molecule to its sequence, using a scheme that is analogous to nucleic acid footprinting. The protein is end labeled radioactively and subjected to limited proteolysis, and the products are analyzed by denaturing polyacrylamide gel electrophoresis and autoradiography. The method is tested with the heme protein myoglobin and applied to mapping the (unknown) surface of the molecule lacking the heme group: apomyoglobin. Sites of protein-protein interaction can be identified, as illustrated by footprinting the association between myoglobin and an anti-myoglobin monoclonal antibody.
Zhong
M
M
Department of Chemistry, New York University, NY 10003.
Lin
L
L
Kallenbach
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NR
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001059:Apoproteins / Q000737:chemistry; D001665:Binding Sites; D002417:Cattle; D003001:Cloning, Molecular; D010450:Endopeptidases; D004926:Escherichia coli; D007553:Isotope Labeling / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D009211:Myoglobin / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D014907:Whales
D000911:Antibodies, Monoclonal; D001059:Apoproteins; D009211:Myoglobin; D010761:Phosphorus Radioisotopes; D011506:Proteins; D011994:Recombinant Proteins; C010857:apomyoglobin; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D010450:Endopeptidases