amino-terminal extension present in the methionine aminopeptidase type 1c of mycobacterium tuberculosis is indispensible for its activity伴扩展中蛋氨酸氨基肽酶1型c的结核分枝杆菌是必不可少的活动.pdfVIP

Kanudia et al. BMC Biochemistry 2011, 12:35 /1471-2091/12/35 RESEARCH ARTICLE Open Access Amino-terminal extension present in the methionine aminopeptidase type 1c of Mycobacterium tuberculosis is indispensible for its activity * Pavitra Kanudia, Monica Mittal, Sangaralingam Kumaran and Pradip K Chakraborti Abstract Background: Methionine aminopeptidase (MetAP) is a ubiquitous enzyme in both prokaryotes and eukaryotes, which catalyzes co-translational removal of N-terminal methionine from elongating polypeptide chains during protein synthesis. It specifically removes the terminal methionine in all organisms, if the penultimate residue is non-bulky and uncharged. The MetAP action for exclusion of N-terminal methionine is mandatory in 50-70% of nascent proteins. Such an activity is required for proper sub cellular localization, additional processing and eventually for the degradation of proteins. Results: We cloned genes encoding two such metalloproteases (MtMetAP1a and MtMetAP1c) present in Mycobacterium tuberculosis and expressed them as histidine-tagged proteins in Escherichia coli. Although they have different substrate preferences, for Met-Ala-Ser, we found, MtMetAP1c had significantly high enzyme turnover rate as opposed to MtMetAP1a. Circular dichroism spectroscopic studies as well as monitoring of enzyme activity indicated high temperature stability (up to 50°C) of MtMetAP1a compared to that of the MtMetAP1c. Modelling of MtMetAP1a based on MtMetAP1c crystal structure revealed the distinct spatial arrangements of identical active site amino acid residues and their mutations affected the enzymatic activities of both the proteins. Strikingly, we observed that 40 amino acid long N-terminal extension of MtMetAP1c,