Elucidation of kinetic and structural properties of eye lens ζ-crystallin: an in vitro and in silico approach

The Arabian Camelus dromedarius contains significant concentration of eye lens zeta-crystallin. This enzyme is also present in other life forms including humans, however in lower catalytic amounts. The recombinant camel zeta-crystallin was expressed in the E. coli BL21 (DE3) pLysS strain and purified using HisTrap column. The K-m of the enzyme for 9,10-phenanthrenequinone (9,10-PQ) substrate and NADPH cofactor was determined to be 11.66 and 50.93 mu M, respectively. The V-max for 9,10-PQ and NADPH was obtained as 23.19 and 19.98 mu M min(-1), respectively. The optimum activity of the purified enzyme was found to be at pH 6.0 and at 55 degrees C. Different physico-chemical parameters were analysed including instability index (II), aliphatic index (AI) and the GRAVY index to establish proper characterization. The sequence of the recombinant zeta-crystallin was subjected to homology modelling using SWISS-MODEL webserver followed by validation of the modelled target structure. The evaluation of the modelled zeta-crystallin was performed by several parameters including Ramachandran plot, Z-score values followed by molecular dynamics (MD) simulation. The cumulative analysis of the physico-chemical, quantitative, qualitative and the essential dynamics of simulation of zeta-crystallin and its complexes with 9,10-PQ and NADPH helped in verifying the acceptable quality and stability of the zeta-crystallin structure. Communicated by Ramaswamy H. Sarma

Automated summary

This study investigated the characteristics and structure of Arabian Camelus dromedarius eye lens zeta-crystallin, analyzed the enzyme kinetics parameters and physico-chemical parameters in 9,10-phenanthrenequinone substrate and NADPH cofactor, and established proper characterization. The structure of the recombinant zeta-crystallin was validated through homology modeling and molecular dynamics simulation.