Structure of human erythrocyte spectrin. I. Isolation of the alpha-I domain and its cyanogen bromide peptides.
The alpha-I domain of human erythrocyte spectrin was produced by a mild tryptic digestion of the intact molecule and purified by a single step affinity chromatography procedure using a monoclonal antibody. A tryptic peptide representing the alpha-I domain, which migrated on polyacrylamide gels as an 80,000-dalton peptide, was subjected to automated Edman-Begg degradation. Products from automated sequencing were identified by reverse-phase high performance liquid chromatography. Two smaller proteolytic products of the alpha-I domain (T74 and T50) were also subjected to automated sequence analysis. CNBr cleavage of the alpha-I domain produced nine unique peptides which were separated by gel filtration on a high performance liquid chromatograph. Peptides were further purified by reverse-phase chromatography and characterized by amino acid analysis. Partial sequences were determined by automated NH2-terminal sequence analysis. A single aspartate-proline bond, which was partially hydrolyzed during the cyanogen bromide cleavage reaction, was also identified. These sequence data include the first 86 residues of the alpha-I domain, and the spectrin oligomer binding site has been tentatively localized within the first 39 residues. The sequence of 293 residues of a total 633 residues in the alpha-I domain is presented and represents the first structural information for this protein.
D000595:Amino Acid Sequence; D003488:Cyanogen Bromide; D004912:Erythrocytes / Q000032:analysis; D006801:Humans; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000302:isolation & purification*; D013049:Spectrin / Q000032:analysis*; D014357:Trypsin / Q000378:metabolism
D046911:Macromolecular Substances; D010446:Peptide Fragments; D013049:Spectrin; D014357:Trypsin; D003488:Cyanogen Bromide