title: Manduca sexta serpin-3 regulates prophenoloxidase activation in response to infection by inhibiting prophenoloxidase-activating proteinases.
abstract: Many serine proteinase inhibitors of the serpin superfamily have evolved in vertebrates and invertebrates to regulate serine proteinase cascades that mediate the host defense responses. We have isolated an immune-responsive serpin from the tobacco hornworm, Manduca sexta. This inhibitor, M. sexta serpin-3, contains a reactive site loop strikingly similar to the proteolytic activation site in prophenoloxidase (pro-PO). Molecular cloning and sequence comparison indicate that serpin-3 is orthologous to Drosophila melanogaster serpin 27A, a regulator of melanization. M. sexta serpin-3 is constitutively present in hemolymph at a low concentration of 5-12 microg/ml and increases to 30-75 microg/ml after a microbial challenge. Recombinant serpin-3 efficiently blocks pro-PO activation in the hemolymph, and it forms SDS-stable acyl-enzyme complexes with purified pro-PO-activating proteinases (PAPs) from M. sexta. PAP-serpin-3 complexes were isolated by immunoaffinity chromatography from hemolymph activated by treatment with Micrococcus luteus. Kinetic analysis of PAP-serpin-3 association strongly suggests that serpin-3 is a physiological regulator of the pro-PO activation reaction.
authors.affiliation: Department of Biochemistry, Kansas State University, Manhattan, KS 66506, USA.
authors.forename: Yifei
authors.initials: Y
authors.lastname: Zhu
authors.forename: Yang
authors.initials: Y
authors.lastname: Wang
authors.forename: Maureen J
authors.initials: MJ
authors.lastname: Gorman
authors.forename: Haobo
authors.initials: H
authors.lastname: Jiang
authors.forename: Michael R
authors.initials: MR
authors.lastname: Kanost
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004156:Catechol Oxidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006458:Hemolymph / Q000737:chemistry; D007239:Infections / Q000276:immunology*; D018402:Manduca / Q000201:enzymology* / Q000378:metabolism; D008969:Molecular Sequence Data; D011480:Protease Inhibitors; D016415:Sequence Alignment; D017421:Sequence Analysis; D015843:Serpins / Q000276:immunology / Q000502:physiology*
chemical_list: D004792:Enzyme Precursors; D011480:Protease Inhibitors; D015843:Serpins; C038451:pro-phenoloxidase; D004156:Catechol Oxidase