Practical Protein Chemistry - A. Darbre 1989

Enzymatic fragmentation of the polypeptide chain
Proteases with low specificity
Papain

Papain is an SH-enzyme isolated from the latex of the melon tree (papaya). Detailed information on The properties of this enzyme can be found in [32]. The enzyme is commercially available from A number of companies.

3.6.4.1. Specificity and Hydrolysis Conditions.

Specificity. The substrate specificity of papain has been studied in detail [85]. It has been shown that the enzyme contains a fairly extended binding site, with the hydrolysis of the specific bond resulting from interaction with one of the fragments of this region. The binding site exhibits a high affinity for two consecutively positioned phenylalanine residues in the substrate [32]. Overall, papain hydrolyzes A wide variety of peptide bonds and, consequently, the outcome of the hydrolysis is practically unpredictable.

Papain has been used to obtain large fragments of native Proteins, such as IMMUNOGLOBULINS [79], Myosin [63], and the extracellular fragment of human Histocompatibility Antigens [80].

Hydrolysis conditions. Papain exhibits maximum activity in the pH range of 5–7.5, and enzyme activation is carried out in the presence of sulfhydryl Reagents. Hydrolysis is performed in 0.2 M pyridine-acetate buffer (pH 6.5) containing 1% (v/v) 2,3-dimercaptopropan-1-ol for 1 h at 37 °C, using an Enzyme-to-substrate ratio of 1:50 (v/v) [84]. Papain is readily inactivated by oxidation in the presence of low concentrations of Cysteine, heavy metal salts, or cyanate ions [97]. Papain is stable in the presence of urea, retaining its activity even in an 8 M urea solution; however, the urea solution must be thoroughly deionized to eliminate the inactivating effect of cyanate ions.