The Production and Isolation of Peptides for Sequence Analysis

Abstract

Although this chapter and Chapter 5 will consider the approaches necessary for determining the total amino acid sequence of a protein, it should be stressed that this is not always the ultimate aim of protein sequence studies. Determination of the total sequence of a protein is both a time-consuming (sometimes taking years) and expensive procedure, and such a project should not be taken upon lightly and without good reason. However, the trend in protein sequence analysis nowadays is not always towards the determination of total primary structures but more the rapid generation of partial sequence data sufficient to complement other observations. In recent years rapid developments in the fields of genetic engineering and DNA sequence analysis have made the direct sequencing of the gene for a given protein (and hence the deduction of its amino acid sequence) a viable alternative, in some cases, to the direct sequencing of a protein. It should be stressed, however, that some protein sequence data is still necessary for complete interpretation of the DNA sequence data. For example, comparison of peptide sequence data with DNA sequence data is necessary to identify initiation, termination and intron coding regions, or indeed the DNA coding region itself. Additionally, DNA sequence data provides no information whatsoever concerning the many protein modifications (e.g. acetylation, methylation, glycosylation, phosphorylation, cleavage of signal or pro-sequences etc.) that are known to occur as post-translational events, and these modifications can only be determined by peptide analysis.