118] Sequence methods

Abstract

Publisher Summary This chapter discusses the developments in the methods available for the determination of amino acid sequences in peptides. The methods include partial hydrolysis with dilute acetic acid, digestion with elastase, digestion with leucine amino peptidase, Sjoquist's modification of the Edman stepwise degradation method, and fingerprinting of mixtures of peptides. Quantitative analysis of most amino acids could be achieved by one-dimensional chromatography on the Whatman No. 1 paper in n -butanol–acetic acid–water, 4:1:5 by volume. A complete separation of the amino acids alanine, aspartic acid, arginine, glutamic acid, glycine, leucine, lysine, and valine, as well as a mixture containing glutamic acid, histidine, threonine, leucine, lysine, proline, and valine, in a manner suitable for quantitative analysis, was achieved by using a modification of the Redfield two-dimensional chromatography system. The denatured protein is digested with trypsin, and the resulting mixture of peptides is characterized by a combination of paper electrophoresis and paper chromatography. Each peptide occupies a position on the final map or fingerprint which is characteristic of its amino acid composition and, to a much smaller degree, of its sequence.