Substrate specificities of growth factor associated kallikreins of the mouse submandibular gland.
The kinetic constants for the hydrolysis of a series of tripeptide p-nitroanilide substrates by mouse epidermal growth factor binding protein (EGF-BP), the gamma-subunit of mouse nerve growth factor (gamma-NGF), bovine pancreatic trypsin (BPT), and porcine pancreatic kallikrein (PPK) have been evaluated. These substrates correspond to the carboxyl-terminal three amino acids of the mature forms of epidermal growth factor (EGF) and beta-nerve growth factor (beta-NGF), as well as various substitutions in the penultimate and antepenultimate positions, and, as such, represent potential recognition sites for precursor processing. The mouse kallikreins (EGF-BP and gamma-NGF) preferentially hydrolyze the substrates with the sequences of their specifically associated growth factors; however, the constants derived from these reactions do not account for the association constants observed with the mature growth factors, and additional significant binding interactions between EGF-BP and EGF and between gamma-NGF and beta-NGF are predicted to exist outside of the catalytic binding site, i.e., the P3 to P1 positions. A comparison of the kinetic constants of BPT, PPK, and the mouse kallikreins indicates that EGF-BP and gamma-NGF display a hybrid catalytic character. A favorable substrate P1 arginine guanidinium group interaction exists for the mouse kallikreins, similar to that of BPT, but a preference for a hydrophobic side chain in the substrate P2 position makes the mouse kallikreins, especially EGF-BP, more closely resemble PPK than BPT. These findings have significant implications with regard to molecular modeling of the mouse kallikreins.
Blaber
M
M
Department of Biological Chemistry, College of Medicine, University of California, Irvine 92717.
Isackson
P J
PJ
Marsters
J C
JC
Burnier
J P
JP
Bradshaw
R A
RA
D000595:Amino Acid Sequence; D000818:Animals; D006133:Growth Substances / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007610:Kallikreins / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D008956:Models, Chemical; D013329:Structure-Activity Relationship; D013363:Submandibular Gland; D013379:Substrate Specificity
D006133:Growth Substances; D007610:Kallikreins