title: Specificity profiling of seven human tissue kallikreins reveals individual subsite preferences.
abstract: Human tissue kallikreins (hKs) form a family of 15 closely related (chymo)trypsin-like serine proteinases. These tissue kallikreins are expressed in a wide range of tissues including the central nervous system, the salivary gland, and endocrine-regulated tissues, such as prostate, breast, or testis, and may have diverse physiological functions. For several tissue kallikreins, a clear correlation has been established between expression and different types of cancer. For example, the prostate-specific antigen (PSA or hK3) serves as tumor marker and is used to monitor therapy response. Using a novel strategy, we have cloned, expressed in Escherichia coli or in insect cells, refolded, activated, and purified the seven human tissue kallikreins hK3/PSA, hK4, hK5, hK6, hK7, hK10, and hK11. Moreover, we have determined their extended substrate specificity for the nonprime side using a positional scanning combinatorial library of tetrapeptide substrates. hK3/PSA and hK7 exhibited a chymotrypsin-like specificity preferring large hydrophobic or polar residues at the P1 position. In contrast, hK4, hK5, and less stringent hK6 displayed a trypsin-like specificity with strong preference for P1-Arg, whereas hK10 and hK11 showed an ambivalent specificity, accepting both basic and large aliphatic P1 residues. The extended substrate specificity profiles are in good agreement with known substrate cleavage sites but also in accord with experimentally solved (hK4, hK6, and hK7) or modeled structures. The specificity profiles may lead to a better understanding of human tissue kallikrein functions and assist in identifying their physiological protein substrates as well as in designing more selective inhibitors.
authors.affiliation: Max-Planck-Institut für Biochemie, Proteinase Research Group, Am Klopferspitz 18, D-82152 Martinsried, Germany.
authors.forename: Mekdes
authors.initials: M
authors.lastname: Debela
authors.forename: Viktor
authors.initials: V
authors.lastname: Magdolen
authors.forename: Norman
authors.initials: N
authors.lastname: Schechter
authors.forename: Martina
authors.initials: M
authors.lastname: Valachova
authors.forename: Friedrich
authors.initials: F
authors.lastname: Lottspeich
authors.forename: Charles S
authors.initials: CS
authors.lastname: Craik
authors.forename: Youngchool
authors.initials: Y
authors.lastname: Choe
authors.forename: Wolfram
authors.initials: W
authors.lastname: Bode
authors.forename: Peter
authors.initials: P
authors.lastname: Goettig
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007313:Insecta; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D020840:Tissue Kallikreins / Q000737:chemistry*; D014357:Trypsin / Q000737:chemistry
chemical_list: D020840:Tissue Kallikreins; D014357:Trypsin