Cloning, overexpression, and purification of bacteial methionine aminopeptidase.

Abstract

Antibiotic resistance has become an important issue in the treatment of microbial diseases in recent years. One approach by the scientific community is the development of novel inhibitors, which target essential bacterial enzymes needed for cell survival. This project focused on methionine aminopeptidase (MetAP), an essential enzyme due to its function of cleaving the first methionine amino acid in premature proteins. In this study, we have cloned and overexpressed MetAP from Bacillus anthracis, Thermotoga maritima, Haemophilus irifluenzae, Staphylococcus aureus, and Oceanobacillus iheyensis as a MetAP-histidine fusion protein. After induction of the gene, MetAP from T. maritima, H irifluenzae, and B. anthracis were successfully purified via nickel affinity chromatography. Enzymatic activity of the proteins, however, could not be reproducibly