Construction protein disuiphide isomerase mutant on saccharomyces cerevisia

Abstract

Protein disulphide isomerase PDI is an enzyme that catalyses disuiphide-bond fonnation in protein to maintain the native conformation, either through redox or isomertzation reactions. Few PDI's have been isolated From various organism such as manunalian liven, plants, green algae and Saceharomyces cerevisiae. Disruption of the PD! in S. cerevisiae is haplo lethal indicating that the product of this gene is essential for viability. PDI consist of 1590 ph, but the essential domain on PD! gene have not been known complete. To study characteristics PDI protein domains, in this research was undertaken construction PD! mutant on S. cerevisiae. !n vitro mutagenesis was carried out using hidroxilamin HA as mutagcn. Recombinant plasnild were constructed by ligation of mutated DNA fragment 758 ph into a vector 6300 ph using fl DNA ligase. The result showed some mutants were lethal, indicated that b and b' domains are essential for PD! on S. cerevisiac. Few mutants poorly growth, and the other showed well growth.