Developments in protein microsequencing.
The author has described here several methods which have been developed in his laboratory. These methods are explained in the following order: [I] gel electrophoresis; 1) extraction from the conventional polyacrylamide gel, 2) a new polyacrylamide gel electrophoresis system for small peptides. [II] Amino acid composition (rapid and micro methods for hydrolysis of proteins); 1) hydrolysis with a mixture of trifluoroacetic acid and hydrochloric acid, 2) vapour hydrolysis, 3) tryptophan micro-analysis. [III] Amino-terminal sequencing; 1) a sensitive detection method of Edman degradation, 2) N-terminal specific labeling method. [IV] Carboxy-terminal sequencing by carboxypeptidase digestion; 1) field desorption mass spectrometry, 2) the use of detergents or alcohols. [V] Monitoring of amino groups at the DNA binding site of protein. As mentioned in the beginning, techniques in the protein field tend to be behind comparable techniques in the DNA field. In addition, the fact that the most important biological molecules presently studied are made in such small amounts in the cell has demanded the development of techniques using precisely and quickly only micro amounts of protein. Such developments of protein chemistry techniques for purification, composition, sequencing, and chemical modification are very much needed in the cell biology field and close collaboration is required between protein chemistry, biophysics, and modern biology in order to develop the above methodology.
Tsugita
A
A
Life Science Institute, Science University of Tokyo, Chiba, Japan.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002268:Carboxypeptidases / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D013058:Mass Spectrometry; D011506:Proteins / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism
D000596:Amino Acids; D011506:Proteins; D004247:DNA; D002268:Carboxypeptidases