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PubMed · 667015

Direct microsequence analysis of polypeptides using an improved sequenator, a nonprotein carrier (polybrene), and high pressure liquid chromatography.

M W Hunkapiller·L E Hood
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Abstract

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 Wittmann-Liebold [Wittmann-Liebold, B. (1973) Hoppe-Seyler's Z. Physiol. Chem. 354, 1415] to analyze amino acid phenylthiohydantoins 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 myoglobin starting with 200 pmol of protein, 77 residues of an antibody light chain with 5 nmole 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, extende N-terminal sequence analysis for nanomole and subnanomole levels of polypeptides and protines, and it also is applicable to analysis of more classical sample quantities.

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Amino Acid SequenceChromatography, High Pressure LiquidHexadimethrine BromideMethodsMicrochemistryMyoglobinPeptidesComparative StudyJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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M W Hunkapiller, L E Hood. 1978-05-30. Direct microsequence analysis of polypeptides using an improved sequenator, a nonprotein carrier (polybrene), and high pressure liquid chromatography.. https://doi.org/10.1021/bi00604a016

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