In vitro PAI-1 can be reactivated by denaturants such as SDS guanidine HCl and urea

protease inhibitor from the hard tick Rhipicephalus appendiculatus that is a potent b-tryptase inhibitor, but not for urokinase, thrombin, factor Xa, factor XIIa, elastases, kallikreins, cathepsin G, granzyme B, chymase and chymotrypsins. Hard tick feeding lasts up to a week as opposed to their distant relative, the soft ticks, whose feeding cycle is much faster. Because of the extended hard tick feeding cycle, a complex of host defense responses takes place at the injury site that is counteracted by the pharmacological properties of tick saliva. Tick salivary protease inhibitors play a role in regulating host proteolytic events and the transmission of tick-borne diseases, such as Lyme disease, while other tick salivary proteins facilitate the transmission of rickettsioses and tick-borne encephalitis. Because of the known pharmacological properties of tick saliva, two salivary gland transcriptome and proteome projects �C also called sialome projects �C revealed secreted salivary proteins expressed from the hard tick, Ixodes scapularis. Annotating these sialome projects amounted to hundreds of tick salivary sequences that remain uncharacterized. These projects revealed many protein sequences classified as having the conserved Kunitz domain and 60 sequences are annotated as monolaris �C sequences that have six cysteine residues forming three disulfide bridges and a single Kunitz head. These 60 monolaris sequences can be further divided into JSI-124 subgroups categorized by variations in their Cys motif. The remaining Kunitz sequences from I. scapularis are defined as bilaris and penthalaris. In our study we focused on the most abundant Kunitz group from the I. scapularis sialome project by Ribeiro et al. : the monolaris group. We identified a Kunitz sequence that displays an Fmoc-Val-Cit-PAB-MMAE unusal Cys motif when compared with the other monolaris and to previously reported Kunitz peptides. Since tick Kunitz peptides are known to inhibit serine proteases we performed an inhibitory screening demonstrating that this I. scapularis Kunitz inhibits several proteases as well as being a potent inhibitor of human skin b-tryptase. Furthermore, a phylogenetic analysis using several functionally described Kunitz protease inhibitors from hematophagous arthropods, nematodes and platyhelminthes