Biochemical characterisation of MX-4, a plant cysteine protease of broad specificity and high stability

A new proteinase, mexicain-IV (MX-4), has been purified to homogeneity from the latex fruits of Jacaratia mexicana (formely Pileus mexicanus), a plant member of the Caricaceae family. MX-4 shows a Mr of 23.7 kDa, pI of 9.3 and maximum proteolytic activity on casein and BAPNA at pH 8.0–8.5 and pH 7.0–7.5, respectively. The amino acid sequence of MX-4 and its reversible inhibition by HgCl2 show that the proteinase belongs to the family of cysteine proteinases. This enzyme exhibits rather broad substrate specificity, although there seems to be a slight preference for cleavage of peptides having certain hydrophobic residues in the P2 position. Biochemical and circular dichroism studies revealed that this enzyme belongs to the α + β class of proteins, in agreement with the results obtained by X-ray crystallographic structure determination, and showed that MX-4 has a higher pH and thermal stability than other members of the Caricaceae family, including papain. These properties make this novel protease a suitable novel analytical tool for the proteomic analysis of peptide fragments of great potential interest in the food industry and other industries.