Manduca sexta serpin-3 regulates prophenoloxidase activation in response to infection by inhibiting prophenoloxidase-activating proteinases.
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.
Zhu
Yifei
Y
Department of Biochemistry, Kansas State University, Manhattan, KS 66506, USA.
Wang
Yang
Y
Gorman
Maureen J
MJ
Jiang
Haobo
H
Kanost
Michael R
MR
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*
D004792:Enzyme Precursors; D011480:Protease Inhibitors; D015843:Serpins; C038451:pro-phenoloxidase; D004156:Catechol Oxidase