Zingibain

Zingibain, zingipain, or ginger protease (EC 3.4.22.67) is a cysteine protease enzyme found in ginger (Zingiber officinale) rhizomes. It catalyses the preferential cleavage of peptides with a proline residue at the P2 position. It has two distinct forms, ginger protease I (GP-I) and ginger protease II (GP-II). Zingibain, zingipain, or ginger protease (EC 3.4.22.67) is a cysteine protease enzyme found in ginger (Zingiber officinale) rhizomes. It catalyses the preferential cleavage of peptides with a proline residue at the P2 position. It has two distinct forms, ginger protease I (GP-I) and ginger protease II (GP-II). As a member of the papain family of cysteine proteases, zingibain shares several structural and functional similarities with more well-studied enzymes such as papain, bromelain, and actinidin. These peptidases contain an active cysteine residue in their centers that catalyzes the hydrolytic cleavage of peptide bonds. Zingibain is noted for its activity as a proteinase and a collagenase. It was first isolated, purified, and reported in 1973 by Ichikawa et al at Japan Women’s University. Recently, zingibain was found to exist as two isozymes, GP-I and GP-II, which were isolated by chromatography, with molecular weights of approximately 22,500 Da. Zingibain utilizes a catalytic triad of Cys, His, and Asn residues in its active site in order to cleave peptide bonds hydrolytically. The presence of Asn175 stabilizes the imidazole ring of His, ensuring it is positioned optimally to catalyze hydrolysis. The mechanism begins with a proton transfer from Cys25 to His159. The sulfide anion then attacks the amino acid's alpha carbon, displacing the amine group, which attaches to His159. The alpha carbon on the stabilized amino acid is then attacked by a water molecule, which displaces the sulfide of Cys25 to convert the amino acid to a carboxylic acid, which is released from the enzyme active site. The experimental introduction of dithiothreitol, a known thiol group protector, improves proteolytic activity, providing further verification of the importance of the central cysteine residue to enzymatic activity. Zingibain exhibits maximum turnover rate at 60°C and rapidly denatures at 70°C. Proteolysis is largely unhampered during cooking with ginger. Optimal temperature ranges of papain and ficin are elevated relative to zingibain, whereas bromelain operates at a slightly lower range. Maximum proteolytic activity of zingibain occurs at pH of 6.0, although the enzyme is still active in pH ranges from 4.5 to 6.0 (optimal pH for meat marinades). GP-II, the more acidic of the two isozymes, exhibits a pI of 4.82, and GP-I exhibits pI values at 5.05 or 5.16. These multiple pI values lend support to a theory that GP-I may be a mixture of two proteins.