Citation

Hermans JM, Monard D, Jones R, Stone SR. Biochemistry 1995; 34(11): 3678-3685.

Copyright

(Copyright © 1995, American Chemical Society)

DOI

10.1021/bi00011a024

PMID

7893664

Abstract

The serpins antithrombin, protease nexin 1, and alpha 1-antitrypsin with a reactive-center arginine (Arg-alpha 1-antitrypsin) were found to inhibit the sperm protease acrosin with varying efficiency. The serpins were titrated against acrosin to determine their specific activity with respect to this enzyme. While antithrombin was fully active against acrosin, more than one molecule of Arg-alpha 1-antitrypsin and protease nexin 1 was required to inhibit one molecule of acrosin. In particular, only 2.7% of protease nexin 1 molecules interacting with acrosin formed stable complexes with the enzyme at 37 degrees C and this value decreased to 0.03% at 12 degrees C. N-terminal sequence analysis indicated that acrosin had cleaved protease nexin 1 at its reactive-center Arg-Ser bond. The results could be interpreted in terms of protease nexin 1 acting as a suicide substrate for acrosin; after the formation of an initial complex, the serpin partitioned between pathways yielding either inactivated (cleaved) serpin or a stable serpin-enzyme complex. The association rate constant (k(ass)) and inhibition constant (Ki) for the stable complexes were determined for each of the serpins by using slow-binding kinetics. The values of k(ass) were 2 x 10(5), 4 x 10(4), and 5 x 10(3) M-1 s-1 for Arg-alpha 1-antitrypsin, antithrombin, and protease nexin 1, respectively. The Ki values for the serpins were 1 nM or less. Heparin markedly accelerated the inhibition of acrosin by antithrombin and protease nexin 1; at the optimal concentration, the degree of heparin acceleration of the inhibition rate was 250- and 500-fold for antithrombin and protease nexin 1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Language: en

Keywords

Acrosin; Amino Acid Sequence; Heparin; Kinetics; Molecular Sequence Data; Serpins; Substrate Specificity