Matrix metalloproteinase-3 (MMP-3, or stromelysin-1) specifically hydr
olyzes the Glu(143)-Leu(144) peptide bond in 45-kDa single-chain uroki
nase-type plasminogen activator (scu-PA) and in its two-chain (tcu-PA)
derivative, yielding a 17-kDa NH2-terminal domain comprising the u-PA
receptor (u-PAR) binding site and a 32-kDa COOH-terminal moiety conta
ining the serine proteinase domain of u-PA. The conversion is complete
ly abolished in the presence of the MMP inhibitors EDTA or 1,10-phenan
throline. Biospecific interaction analysis indicates that binding of M
MP-3 occurs through the 32-kDa fragment. The 32-kDa fragment derived f
rom scu-PA (scu-PA-32k) has a specific activity of less than or equal
to 500 IU/mg, but it can be activated with plasmin to a two-chain deri
vative (tcu-PA-32k) with a specific activity of 79 000 IU/mg, tcu-PA a
nd tcu-PA-32k moieties derived from scu-PA-32k by plasmin or from tcu-
PA by MMP-3 have comparable amidolytic activities toward the chromogen
ic substrate S-2444 (k(cat)/K-m of 110 and 160 mM(-1) s(-1), respectiv
ely) and similar plasminogen activating activities in a coupled chromo
genic substrate assay. Specific binding of the 17-kDa NH2-terminal dom
ain to THP-1 monocytoid cells is completely abolished by competition w
ith scu-PA but is not affected by scu-PA-32k (residual binding of 88 /- 9% (mean +/- SEM; n = 3) with 25-fold molar excess). Thus, MMP-3 re
moves a functional NH2-terminal u-PAR-binding domain from u-PA without
affecting its enzymatic properties.