Method for purification of large cyanogen bromide peptides by carboxymethyl cellulose chromatography in 8 m urea: Application to the purification of cyanogen bromide peptides from staphylcoccal enterotoxin A, phosphoglycerate kinase, and glucose-6-phosphate dehydrogenase
✍ Scribed by I-Yih Huang; Carol Ewing; Akira Yoshida
- Publisher
- Elsevier Science
- Year
- 1976
- Tongue
- English
- Weight
- 399 KB
- Volume
- 73
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
A method for purification of large cyanogen bromide peptides from proteins by means of carboxymethyl cellulose chromatography in the presence of 8 M urea is described. Chromatography ofa number of large cyanogen bromide peptides which could not be separated by gel filtration showed that the resolution of the system was sufficient to enable large cyanogen bromide peptides to be separated from one another. The use of this method to purify cyanogen bromide peptides of a protein as a first step is also discussed.
For the elucidation of the primary structure of proteins, the digestion of the protein with cyanogen bromide (1) has become a popular procedure. Because of its specificity for methionyl residues and the fact that methionine is usually rare in proteins, the cyanogen bromide cleavage of proteins causes the formation of large peptides which provide the essential information concerning the correct positioning of peptides derived from enzymatic digests or the confirmation of a previously established sequence. Since an automated sequence analyzer can be efficiently applied to larger peptides. extensive information on the protein structure will be obtained by analyzing large cyanogen bromide peptides.
Thus, the separation of peptide mixtures obtained from the cyanogen bromide cleavage of a protein has become very important. Molecular sieve chromatography has commonly been used as a first step to separate cyanogen bromide peptide mixtures, and recyclings on the same systems to purify these peptides as the second step have been widely employed (2-6). Preparative polyacrylamide-gel electrophoresis at pH 8.9 in 8 M urea (7), diethylaminoethyl cellulose (8), diethylaminoethyl Sephadex (9),