title: Manduca sexta serpin-7, a putative regulator of hemolymph prophenoloxidase activation.
abstract: Serpins regulate various physiological reactions in humans and insects, including certain immune responses, primarily through inhibition of serine proteases. Six serpins have previously been identified and characterized in the tobacco hornworm Manduca sexta. In this study, we obtained a full-length cDNA sequence of another Manduca serpin, named serpin-7. The open reading frame of serpin-7 encodes a polypeptide of 400 amino acid residues with a predicted signal peptide of the first 15 residues. Multiple protein sequence alignment of the reactive center loop region of the M. sexta serpins indicated that serpin-7 contains Arg-Ile at the position of the predicted scissile bond cleaved by protease in the serpin inhibition mechanism. The same residues occur in the scissile bond of the reactive center loop in M. sexta serpin-4 and serpin-5, which are protease inhibitors that can block prophenoloxidase activation in plasma. Serpin-7 transcript was detected in hemocytes and fat body, and its expression increased in fat body after injection of larvae with Micrococcus luteus. Recombinant serpin-7 added to larval plasma inhibited spontaneous melanization and decreased prophenoloxidase activation stimulated by bacteria. Serpin-7 inhibited prophenoloxidase-activating protease-3 (PAP3), forming a stable serpin-protease complex. Considering that serpin-3 and serpin-6 are also efficient inhibitors of PAP3, it appears that multiple serpins present in plasma may have redundant or overlapping functions. We conclude that serpin-7 has serine protease inhibitory activity and is likely involved in regulation of proPO activation or other protease-mediated aspects of innate immunity in M. sexta.
authors.affiliation: Department of Biochemistry, Kansas State University, 141 Chalmers Hall, Manhattan, KS 66506, USA.
authors.forename: Chansak
authors.initials: C
authors.lastname: Suwanchaichinda
authors.forename: Rose
authors.initials: R
authors.lastname: Ochieng
authors.forename: Shufei
authors.initials: S
authors.lastname: Zhuang
authors.forename: Michael R
authors.initials: MR
authors.lastname: Kanost
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004156:Catechol Oxidase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006458:Hemolymph / Q000737:chemistry / Q000201:enzymology*; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018402:Manduca / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D015843:Serpins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D019476:Insect Proteins; D015843:Serpins; C038451:pro-phenoloxidase; D004156:Catechol Oxidase