Biochemical and structural analysis of a site directed mutant of manganese dependent aminopeptidase P from Streptomyces lavendulae

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dc.contributor.authorNandan, Arya
dc.contributor.authorNampoothiri, Kesavan M.
dc.date.accessioned2024-05-28T18:51:20Z
dc.date.available2024-05-28T18:51:20Z
dc.date.issued2015-03-19
dc.description.abstractAminopeptidase P (APP) removes N-terminal amino acids from peptides and proteins when the penultimate residue is proline. To understand the structure-function relationships of aminopeptidase P of Streptomyces lavendulae, a conserved arginine residue was replaced with lysine (R453K) by site-directed mutagenesis. The overexpressed wild and mutant enzymes were of nearly 60 kDa and purified by nickel affinity chromatography. Kinetic analysis of R453K variant using Gly-Pro-pNA as the substrate revealed an increase in Km with a decrease in Vmax, leading to overall decrease in the catalytic efficiency, indicating that the guanidinium group of arginine plays an important role in substrate binding in APP. We constructed three dimensional models for the catalytic domains of wild and mutant enzyme and it revealed an interaction in R453 of native enzyme through hydrogen bonding with the adjacent residues making a substrate binding cavity whereas K453 did not participate in any hydrogen bonding. Hence, R453 in APP of S. lavenduale must be playing a critical role in the hydrolysis of the substrate.
dc.identifier.issn1314-6246
dc.identifier.urihttps://doi.uni-plovdiv.bg/handle/store/124
dc.language.isoen
dc.publisherPlovdiv University Press “Paisii Hilendarski”
dc.subjectAminopeptidase P
dc.subjectcatalytic domain
dc.subjectmetalloprotease
dc.subjectsite directed mutagenesis
dc.subjectStreptomyces lavendulae
dc.titleBiochemical and structural analysis of a site directed mutant of manganese dependent aminopeptidase P from Streptomyces lavendulae
dc.typeArticle
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