title: Purification of a serine protease and evidence for a protein C activator from the saliva of the tick, Ixodes scapularis.
abstract: The saliva of ticks is critical to their survival as parasites and hematophagous animals. In this study, we have purified an enzyme with trypsin-like activity from the saliva of the tick vector of Lyme Disease, Ixodes scapularis. This enzyme, named as IXOSP (I. scapularis salivary serine protease), is a 29.9 kDa molecule with N-terminus FPxMVxLRIKxR. A BLAST search identified IXOSP as a secreted serine protease (AAY66740) with a conserved catalytic triad His, Asp, and Ser. In vitro studies demonstrated that IXOSP cleaves chromogenic substrates with arginine in the P1 position, by a mechanism inhibited by PMSF or aprotinin. Gene expression studies revealed that IXOSP is expressed at different tick developmental stages, including eggs, and unfed or fed adult tick salivary glands, but not in nymphs or in the midgut. While the physiological substrate for IXOSP remains to be identified, we demonstrated that I. scapularis saliva activate protein C (PC) resulting in the production of activated PC, a potent anticoagulant that also regulates a myriad of inflammatory responses through protease activated receptors. In contrast, the salivary glands of Anopheles gambiae, Anopheles stephensi, Anopheles albimanus, Aedes aegypti, Lutzomyia longipalpis, and Phlebotomus ariasi did not activate protein C. These discoveries are discussed in the context of blood coagulation, inflammation and vector-host interactions.
authors.affiliation: Center for Vector-Borne Disease, University of Rhode Island, Kingston, RI 02881, USA. Electronic address: au_sudha@yahoo.com.
authors.forename: Sivakamasundari
authors.initials: S
authors.lastname: Pichu
authors.affiliation: Section of Vector Biology, Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: José M C
authors.initials: JM
authors.lastname: Ribeiro
authors.affiliation: Center for Vector-Borne Disease, University of Rhode Island, Kingston, RI 02881, USA.
authors.forename: Thomas N
authors.initials: TN
authors.lastname: Mather
authors.affiliation: Section of Vector Biology, Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA. Electronic address: ifrancischetti@niaid.nih.gov.
authors.forename: Ivo M B
authors.initials: IM
authors.lastname: Francischetti
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D006487:Hemostasis / Q000187:drug effects*; D018884:Ixodes / Q000201:enzymology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000235:genetics / Q000302:isolation & purification* / Q000633:toxicity; D012237:Rhode Island; D012463:Saliva / Q000201:enzymology*; D020539:Sequence Analysis, Protein; D057057:Serine Proteases / Q000235:genetics / Q000302:isolation & purification* / Q000633:toxicity; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D009842:Oligopeptides; C047135:protein C activator peptide; D057057:Serine Proteases
keywords: Activated protein C (APC); Ixodes scapularis; Protease activated receptors (PAR); Saliva; Serine protease