DIRECT MICROSEQUENCE ANALYSIS OF POLYPEPTIDES USING AN IMPROVED SEQUENATOR, A NON-PROTEIN CARRIER (POLYBRENE), AND HIGH-PRESSURE LIQUID-CHROMATOGRAPHY

We have combined the use of a nonprotein carrier (Polybrene), high pressure liquid chromatography, and modifications in Edman chemistry with the improvements of a commercial spinning cup sequenator suggested by Witt-mann-Liebold [Wittmann-Liebold, B. (1973) Hoppe-Seyler's Z. Physiol. Chem. 354, 1415] to analyze amino acid phenyl-thiohydantoins obtained from automated Edman degradation of microquantities of polypeptide directly without the use of radiolabel. This approach has allowed us to determine the sequence of the N-terminal 47 residues of sperm whale myo-globin starting with 200 pmol of protein, 77 residues of an antibody light chain with 5 nmol of protein, and 54 residues of an antibody heavy chain with 8 nmol of protein. In addition, we completely sequenced a hydrophobic 14-residue peptide at the 1.5-nmol level. Our technique of direct analysis for microsamples is capable of providing routine, extended N-terminal sequence analysis for nanomole and subnanomole levels of polypeptides and proteins, and it also is applicable to analysis of more classical sample quantities. © 1978, American Chemical Society. All rights reserved.