PAI-1 NBD-P9 mutant is created via mutagenesis of the P-9 serine residue (Ser338) on the reactive center loop to cysteine. This promoted the availabilty of a free thiol group for incorporation of NBD, a fluorescent probe highly sensitive to changes in dissolution and hydrophobic environment. The fluorescence emission of PAI-1 NBD-P9 is enhanced 6-7 fold upon insertion of the reactive center loop into beta-sheet A following complex formation with proteases, formation of the latent species, or cleavage by elastase. This modified PAI-1 is nearly as active as wild type PAI-1 and is more resistant to the spontaneous latency reaction making it an excellent tool for monitoring reaction rates of PAI-1 (1). PAI-1 NBD-P9 has been utilized in a number of studies to determine the rates of loop insertion and SERPIN reaction mechanisms when reacted with various proteinases (1,2), inactivating antibodies (3) and conformational changes imposed by the binding of vitronectin (4).

References
1. Shore J.D. et al.(1995) J Biol Chem 270:5395-5398.
2. Lawrence D.A. et al.(2000) J Biol Chem 275:5839-5844.
3. Verhamme I.M. et al.(1999) J Biol Chem 274:17511-17517.
4. Gibson A. et al.(1997) J Biol Chem 272:5112-5121.