Enzymatic Cleavage of Proteins

Abstract

Endoproteinases catalyse hydrolysis of polypeptide chains, most usefully at specific sites within the polypeptide, as described in Table 1. The number and nature of peptides generated by a proteinase of good specificity is characteristic of a protein, since it reflects the protein’s sequence. The term & peptide map & is applied to the chromatogram or pattern of peptides resolved by a method such as high-performance liquid chromatography (HPLC) or capillary electrophoresis (see Chapter 8). Peptide mapping is widely used for quality control of recombinant proteins, where appearance of novel peptides indicates the presence of variant forms of protein (for example, see refs. 1–4). The mass spectroscopic equivalent of peptide mapping is called & mass mapping &, whereby the masses of the products of proteolysis are characteristic of a given protein (see Chapters 17 and 18). Individual peptides may also be purified and subjected to various sequencing techniques as described elsewhere in this volume, the purpose being to identify a protein by its sequence, determine the partial sequence of a novel protein for cloning purposes, or identify sites of modification (for example, phosphorylation 5).

Table 1 Characteristics of “Sequencer-Grade” Endoproteases