Identification of plasma proteinase complexes with serpin-3 in Manduca sexta
Autor(es): Christen, Jayne M.; Hiromasa, Yasuaki; An, Chunju; Kanost, Michael R.
Resumo: Extracellular serine proteinase cascades stimulate prophenoloxidase (proPO) activation and antimicrobial peptide production in insect innate immune responses. Serpins in plasma regulate such cascades by selective inhibition of proteinases, in reactions which result in the formation of covalent serpin-proteinase complexes. We carried out experiments to identify plasma proteinases that are inhibited by Manduca sexta serpin-3, an immune-inducible serpin known to regulate proPO activation. Immunoaffinity chromatography, using antiserum to serpin-3, yielded serpin-3 complexes with proteinases identified by immunoblot analysis as prophenoloxidase-activating proteinase (PAP)-1, PAP-2, PAP-3, and hemolymph proteinase 8 (HP8). HP8 can cleave and activate the Toll ligand, Spatzle, leading to synthesis of antimicrobial Peptides. Analysis by mass spectrometry of tryptic Peptides derived from the serpin-3 complexes confirmed the presence of PAP-1. PAP-3, and HP8. Purified recombinant serpin-3 and active HP8 formed an SDS-stable complex in vitro. Identification of serpin-3-proteinase complexes in plasma provides insight into proteinase targets of serpin-3 and extends the understanding of serpin/proteinase function in the immune response of M. sexta. (C) 2012 Elsevier Ltd. All rights reserved.
Palavras-Chave: Serpin; Hemolymph; Immunity; Hemolymph proteinases; Proteomics
Imprenta: Insect Biochemistry and Molecular Biology, v. 42, n. 12, p. 946-955, 2012
Identificador do objeto digital: 10.1016/j.ibmb.2012.09.008
Descritores: Aedes aegypti - Immune response ; Aedes aegypti - Molecular structure ; Aedes aegypti - Protein synthesis
Data de publicação: 2012
