Biochemical characterization of Anopheles gambiae SRPN6, a malaria parasite invasion marker in mosquitoes

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Show simple item record An, Chunju Hiromasa, Yasuaki Zhang, Xin Lovell, Scott Zolkiewski, Michal Tomich, John M. Michel, Kristin 2012-12-20T20:26:28Z 2012-12-20T20:26:28Z 2012-12-20
dc.description.abstract Serine proteinase inhibitors of the serpin family are well known as negative regulators of hemostasis, thrombolysis and innate immune responses. Additionally, non-inhibitory serpins serve functions as chaperones, hormone transporters, or antiangiogenic factors. In the African malaria mosquito, Anopheles gambiae s.s., at least three serpins (SRPNs) are implicated in the innate immune response against malaria parasites. Based on reverse genetic and cell biological analyses, AgSRPN6 limits parasite numbers and transmission and has been postulated to control melanization and complement function in mosquitoes. This study aimed to characterize AgSRPN6 biophysically and determine its biochemical mode of action. The structure model of AgSRPN6, as predicted by I-Tasser showed the protein in the native serpin fold, with three central β-sheets, nine surrounding α-helices, and a protruding reactive center loop. This structure is in agreement with biophysical and functional data obtained from recombinant (r) AgSRPN6, produced in Escherichia coli. The physical properties of purified rAgSRPN6 were investigated by means of analytical ultracentrifugation, circular dichroism, and differential scanning calorimetry tools. The recombinant protein exists predominantly as a monomer in solution, is composed of a mixture of ahelices and b-sheets, and has a mid-point unfolding temperature of 56uC. Recombinant AgSRPN6 strongly inhibited porcine pancreatic kallikrein and to a lesser extent bovine pancreatic trypsin in vitro. Furthermore, rAgSRPN6 formed inhibitory, SDSstable, higher molecular weight complexes with prophenoloxidase-activating proteinase (PAP)1, PAP3, and Hemolymph protein (HP)6, which are required for melanization in the lepidopteran model organism, Manduca sexta. Taken together, our results strongly suggest that AgSRPN6 takes on a native serpin fold and is an inhibitor of trypsin-like serine proteinases. en_US
dc.language.iso en_US en_US
dc.relation.uri en_US
dc.subject Anopheles gambiae en_US
dc.subject Mosquitoes en_US
dc.subject Serpins en_US
dc.subject Malaria parasites en_US
dc.subject AgSRPN6 en_US
dc.title Biochemical characterization of Anopheles gambiae SRPN6, a malaria parasite invasion marker in mosquitoes en_US
dc.type Article (publisher version) en_US 2012 en_US
dc.citation.doi 10.1371/journal.pone.0048689 en_US
dc.citation.epage e48689-9 en_US
dc.citation.issue 11 en_US
dc.citation.jtitle PLoS ONE en_US
dc.citation.spage e48689-1 en_US
dc.citation.volume 7 en_US
dc.contributor.authoreid michalz en_US
dc.contributor.authoreid jtomich en_US
dc.contributor.authoreid kmichel en_US
dc.contributor.authoreid hiromasa en_US
dc.contributor.authoreid xz65 en_US

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