Crystallography & Molecular Biology Division, Saha Institute of Nuclear Physics
○Debi Choudhury Sampa Biswas Chandana Chakrabarti J.K. Dattagupta
Proteases constitute one of the important groups of industrial enzymes. Amongst the proteases, the cysteine proteases are of considerable commercial importance due to their strong proteolytic activity against a broad range of substrates. The widely studied member of this group is Papain, which has immense industrial and pharmaceutical applications, for which it is desirable that papain has higher stability. High stability is of economic advantage because of reduced enzyme turnover, higher reaction rates, and lesser microbial contamination. The kinetic stability of many proteins has been improved by rational design. One strategy for identifying stabilizing mutations involves comparison of more stable homologous proteins with less-stable ones.
In our laboratory we have solved the three dimensional structures of a few naturally occurring highly stable plant cysteine proteases of papain-family. Based on these three dimensional structures and amino acid sequence patterns we have attempted to design a papain molecule, which will have enhanced stability with negligible compromise in its catalytic activity. For this, the 40 kDa wild type papain precursor has been cloned, overexpressed in E. Coli as inclusion bodies, purified and refolded. Site-directed mutagenesis attempts are in progress on this recombinant papain to obtain the desired stability.