Protein Sequence Determination Research Articles

Studies towards the complete sequence determination of proteins by mass spectrometry: Derivatisation of methionine, cysteine and arginine containing peptides

A number of methionine, pyrimidyl ornithine, and two carboxymethyl cysteine containing peptides have given easily interpretable mass spectra following acetylation, and permethylation using a reaction time of about 60 seconds. Using this procedure involatile “salt” formation was prevented in all cases. For arginine containing peptides, refined methods are described for producing high yields of the required derivative in both dicarbonyl condensation and hydrazinolysis reactions. In the latter case in particular, a number of acidic peptides have now been derivatised without extensive peptide bond cleavage and their sequences determined.

Studies towards the complete sequence determination of proteins by mass spectrometry; a rapid procedure for the successful permethylation of histidine containing peptides

It is now widely recognised that mass spectrometric sequencing, involving the use of Nacetyl permethylated peptide derivatives, offers a viable alternative to classical approaches to the primary structure determination of proteins. The method does not as yet enjoy unanimous support, and critics may point to two areas of weakness. The first of these involves strategy. Comparison of the classical Dansyl-Edman procedure [l] with the mass spectrometric approach to the sequenching of individual peptides shows that, with a few exceptions (e.g. peptides with blocked N-termini), mass spectrometry offers no significant advantages in time-saving or sensitivity. Recently, however, we have described a strategy (mixture analysis) [2] which offers the distinct advantage of rapid sequence assignment, preempting the necessity for purification and isolation of individual peptides. The second, more fundamental weakness is associated with the derivatisation of the peptides, in particular the permethylation reaction. The best permethylation procedure to date has been the Hakamori reaction [3], first described for use on peptides by Vilkas and Lederer [4]. A modified procedure described by Thomas [5] is now commonly used, but this does not give satisfactory results with certain of the amino acids normally found in proteins. One of the troublesome amino acids is histidine, which is believed to form a quaternised salt on permethylation [6]. Using the Thomas procedure [5,7] one can at best determine the sequence up to but not including the histidine, and more often no sequence information is obtained at all. This report describes some experiments which have

A manual method of sequential edman degradation followed by dansylation for the determination of protein sequences

A modification of the manual method of sequential Edman degradation followed by dansylation for sequencing peptides has been developed for use on long polypeptides and intact chains of proteins. This method permits the sequence of 15 to 20 residues from the amino-terminal end of the chain to be determined.

Protein Sequence Determination. A Sourcebook of Methods and Techniques. Molecular Biology Biochemistry and Biophysics, Volume 8.Saul B. Needleman

<i>Protein Sequence Determination. A Sourcebook of Methods and Techniques. Molecular Biology Biochemistry and Biophysics, Volume 8.</i>Saul B. Needleman

Protein Sequence Determination. Methods and Techniques.S. Blackburn

Protein Sequence Determination. Methods and Techniques.S. Blackburn

A solid‐phase Edman sequence determination

AbstractThe sequence of the amino acids in a peptide covalently bound to a polymer could be determined by reaction with methyl isothiocyanate. This extension of the Edman degradation is likely to facilitate the automation of the sequence determination of peptides and proteins.

Computer aids to protein sequence determination

Computer programs are described to aid in determining the sequence of the amino acids in a protein from knowledge of the structure of overlapping peptides obtained by hydrolysis of the protein. The programs are designed for use in planning experiments, locating experimental errors, determining all possible sequences consistent with the experimental data and testing the reliability of the final answer. The system of Fortran computer programs accepts the experimental peptide structure data and presents results in acceptable chemical language. Examples are described utilizing different types of experimental information for partially or completely determining the sequence. The possibility of accurate systematic manipulation and presentation of data makes attractive a much wider range of experimental techniques than are now used.