Comparison of human and porcine tissue kallikrein substrate specificities.
Little is known about the species specificity of tissue kallikrein-kininogen interaction since the kinetic parameters for Lys-bradykinin release from kininogen by tissue kallikreins from different animal species have not been reported. We have now determined the kinetic parameters for hydrolysis by human and porcine tissue kallikrein, hK1 and pK1, respectively (Berg et al., 1992) of two series of intramolecularly quenched fluorogenic peptides having the sequences that flank the scissile Arg-Ser or Met-Lys bond in human and bovine kininogen. Results have shown that peptides having sequences from human kininogen are better substrates for hK1 and peptides derived from bovine kininogen are better substrates for pK1. Kinetic data for hydrolysis of the Arg-Ser bond showed that differences in the interaction of residue(s) in positions P2'-P10' contribute to the efficiency of the cleavage and may be responsible for differences in their susceptibilities to the two kallikreins. Significant variations in the kinetic data were observed for the hydrolysis of the Met-Lys bond in substrates with an N-terminal extension at sites P3-P9. The highest k(cat)/Km value in the hydrolysis of Abz-[Gln370-Gln381]-bkng-EDDnp by pk1 demonstrates an important interaction of subsites S5-S4 with Gln and Thr residues in the bovine kininogen segment. A Gln370-Gln391 bovine kininogen fragment used to study the cleavage of both Met-Lys and Arg-Ser bonds in the same molecule confirmed the importance of an extended interaction site for species specificity among tissue kallikreins.
Del Nery
E
E
Universidade Federal de São Paulo-UNIFESP, Escola Paulista de Medicina, Department of Biophysics, Brazil.
Chagas
J R
JR
Juliano
M A
MA
Juliano
L
L
Prado
E S
ES
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000378:metabolism; D002417:Cattle; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D008239:Lysine / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D013045:Species Specificity; D013379:Substrate Specificity; D013552:Swine; D020840:Tissue Kallikreins / Q000378:metabolism*
D005456:Fluorescent Dyes; D009842:Oligopeptides; D010446:Peptide Fragments; D012694:Serine; D001120:Arginine; D008715:Methionine; D020840:Tissue Kallikreins; D008239:Lysine