Molecular cloning, expression and characterization of three distinctive genes encoding methionine aminopeptidases in cyanobacterium Synechocystis sp. strain PCC6803.
Methionine aminopeptidase, known to be encoded by single genes in prokaryotes, is a cobalt-dependent enzyme that catalyzes the removal of N-terminal methionine residues from nascent polypeptides. Three ORFs encoding putative methionine aminopeptidases from the genome of cyanobacterium Synechocystis sp. strain PCC6803, designated as slr0786 ( map-1), slr0918 ( map-2) and sll0555 ( map-3) were cloned and expressed in Escherichia coli. The purified recombinant proteins encoded by map-1 and map-3 had much higher methionine aminopeptidase activity than the recombinant protein encoded by map-2. Comparative analysis revealed that the three recombinant enzymes differed in their substrate specificity, divalent ion requirement, pH, and temperature optima. The broad activities of the iso-enzymes are discussed in light of the structural similarities with other peptidase families and their levels of specificity in the cell. Potential application of cyanobacterial MetAPs in the production of recombinant proteins used in medicine is proposed. This is the first report of a prokaryote harboring multiple methionine aminopeptidases.
Atanassova
Anelia
A
Institute of Molecular Biology, Bulgarian Academy of Sciences, bl. 21 Acad. G. Bonchev St., 1113, Sofia, Bulgaria. atanasso@chem.utoronto.ca
Sugita
Mamoru
M
Sugiura
Masahiro
M
Pajpanova
Tamara
T
Ivanov
Ivan
I
D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism; D003001:Cloning, Molecular; D003035:Cobalt / Q000378:metabolism; D000458:Cyanobacteria / Q000201:enzymology* / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D063208:Methionyl Aminopeptidases; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature
D002413:Cations, Divalent; D007527:Isoenzymes; D011994:Recombinant Proteins; D003035:Cobalt; D000626:Aminopeptidases; D063208:Methionyl Aminopeptidases