title: Human kallikrein 6 activity is regulated via an autoproteolytic mechanism of activation/inactivation.
abstract: Human kallikrein 6 (protease M/zyme/neurosin) is a serine protease that has been suggested to be a serum biomarker for ovarian cancer and may also be involved in pathologies of the CNS. The precursor form of human kallikrein 6 (pro-hK6) was overexpressed in Pichia pastoris and found to be autoprocessed to an active but unstable mature enzyme that subsequently yielded the inactive, self-cleavage product, hK6 (D81-K244). Site-directed mutagenesis was used to investigate the basis for the intrinsic catalytic activity and the activation mechanism of pro-hK6. A single substitution R80 --> Q stabilized the activity of the mature enzyme, while substitution of the active site serine (S197 --> A) resulted in complete loss of hK6 proteolytic activity and facilitated protein production. Our data suggest that the enzymatic activity of hK6 is regulated by an autoactivation/autoinactivation mechanism. Mature hK6 displayed a trypsin-like activity against synthetic substrates and human plasminogen was identified as a putative physiological substrate for hK6, as specific cleavage at the plasminogen internal bond S460-V461 resulted in the generation of angiostatin, an endogenous inhibitor of angiogenesis and metastatic growth.
authors.affiliation: Departament de Bioquímica i Biologia Molecular and Institut de Biotecnologia i Biomedicina, Universitat Autònoma de Barcelona, E-08193 Bellaterra, Spain.
authors.forename: Alex
authors.initials: A
authors.lastname: Bayés
authors.forename: Theodoros
authors.initials: T
authors.lastname: Tsetsenis
authors.forename: Salvador
authors.initials: S
authors.lastname: Ventura
authors.forename: Josep
authors.initials: J
authors.lastname: Vendrell
authors.forename: Francesc X
authors.initials: FX
authors.lastname: Aviles
authors.forename: Georgia
authors.initials: G
authors.lastname: Sotiropoulou
mesh_terms: D004789:Enzyme Activation / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007610:Kallikreins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology
chemical_list: D011994:Recombinant Proteins; D015842:Serine Proteinase Inhibitors; C103551:KLK6 protein, human; D007610:Kallikreins