Irreversible inhibition of serine proteases by peptide derivatives of (alpha-aminoalkyl)phosphonate diphenyl esters.
Peptidyl derivatives of diphenyl (alpha-aminoalkyl)phosphonates have been synthesized and are effective and specific inhibitors of serine proteases at low concentrations. Z-PheP(OPh)2 irreversibly reacts with chymotrypsin (kobsd/[I] = 1200 M-1 s-1) and does not react with two elastases. The best inhibitor for most chymotrypsin-like enzymes including bovine chymotrypsin, cathepsin G, and rat mast cell protease II is the tripeptide Suc-Val-Pro-PheP(OPh)2 which corresponds to the sequence of an excellent p-nitroanilide substrate for several chymases. The valine derivative Z-ValP(OPh)2 is specific for elastases and reacts with human leukocyte elastase (HLE, 280 M-1 s-1) but not with chymotrypsin. The tripeptide Boc-Val-Pro-ValP(OPh)2, which has a sequence found in a good trifluoromethyl ketone inhibitor of HLE, is the best inhibitor for HLE (kobsd/[I] = 27,000 M-1 s-1) and porcine pancreatic elastase (PPE, kobsd/[I] = 11,000 M-1 s-1). The rates of inactivation of chymotrypsin by MeO-Suc-Ala-Ala-Pro-PheP(OPh)2 and PPE and HLE by MeO-Suc-Ala-Ala-Pro-ValP(OPh)2 were decreased 2-5-fold in the presence of the corresponding substrate, which demonstrates active site involvement. Only one of two diastereomers of Suc-Val-Pro-PheP(OPh)2 reacts with chymotrypsin (146,000 M-1 s-1), and the enzyme-inhibitor complex had one broad signal at 25.98 ppm in the 31P NMR spectrum corresponding to the Ser-195 phosphonate ester. Phosphonylated serine proteases are extremely stable since the half-time for reactivation was greater than 48 h for the inhibited elastases and 7.5-26 h for chymotrypsin.(ABSTRACT TRUNCATED AT 250 WORDS)
Oleksyszyn
J
J
School of Chemistry, Georgia Institute of Technology, Atlanta 30332.
Powers
J C
JC
D000818:Animals; D001769:Blood; D002021:Buffers; D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors; D002417:Cattle; D002800:Cholinesterase Inhibitors; D053818:Chymases; D002918:Chymotrypsin / Q000037:antagonists & inhibitors; D005978:Glutathione / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D009842:Oligopeptides / Q000494:pharmacology*; D009943:Organophosphorus Compounds / Q000494:pharmacology*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors; D010455:Peptides / Q000737:chemistry; D011480:Protease Inhibitors / Q000737:chemistry*; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013552:Swine
D002021:Buffers; D002800:Cholinesterase Inhibitors; D009842:Oligopeptides; D009943:Organophosphorus Compounds; D010455:Peptides; D011480:Protease Inhibitors; D015842:Serine Proteinase Inhibitors; D002403:Cathepsins; D012697:Serine Endopeptidases; D002918:Chymotrypsin; C534094:CTSG protein, human; D056649:Cathepsin G; C534097:Ctsg protein, rat; D010196:Pancreatic Elastase; D053818:Chymases; D005978:Glutathione