title: A novel method for selective isolation of C-terminal peptides from tryptic digests of proteins by immobilized anhydrotrypsin: application to structural analyses of the tail sheath and tube proteins from bacteriophage T4.
abstract: A novel method useful for selective isolation of the C-terminal peptide from a tryptic digestion mixture of a protein has been developed by taking advantage of a unique property of anhydrotrypsin, which has a strong specific affinity for the peptides containing arginine or lysine at their C-termini. Briefly, peptides produced by tryptic digestion of a protein are fractionated by affinity chromatography on a column of immobilized anhydrotrypsin. The C-terminal peptide is recovered in a breakthrough fraction, while the remainders are adsorbed on the column (unless the protein ends in arginine or lysine). The breakthrough fraction is then subjected to reversed-phase high-performance liquid chromatography in order to purify the C-terminal peptide. Using this method, we have successfully isolated the C-terminal peptides from tryptic digests of the sheath protein (gp 18) and the tube protein (gp 19) of bacteriophage T4. The analytical results on these peptides, together with the information on the N-terminal structures of the original proteins and on the nucleotide sequences of genes 18 and 19, allowed us to establish the complete primary structures of the two proteins.
authors.affiliation: Department of Biochemistry, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Kumazaki
authors.forename: T
authors.initials: T
authors.lastname: Nakako
authors.forename: F
authors.initials: F
authors.lastname: Arisaka
authors.forename: S
authors.initials: S
authors.lastname: Ishii
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002846:Chromatography, Affinity; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000302:isolation & purification*; D013604:T-Phages / Q000032:analysis*; D014357:Trypsin; D014764:Viral Proteins / Q000302:isolation & purification*
chemical_list: D000596:Amino Acids; D010446:Peptide Fragments; D014764:Viral Proteins; C011661:anhydrotrypsin; D014357:Trypsin