Inhibition of the chymotrypsin-like activity of the pituitary multicatalytic proteinase complex.
The multicatalytic proteinase complex (MPC), also referred to as proteasome, is a large molecular mass intracellular particle (approximately 700 kDa), which exhibits three distinct proteolytic activities designated as chymotrypsin-like, trypsin-like, and peptidylglutamyl-peptide hydrolyzing (PGPH), all sensitive to inhibition by 3,4-dichloroisocoumarin (DCI). The presence of a component resistant to inhibition by DCI with an apparent preference toward bonds on the carboxyl side of branched-chain amino acids has also been recently established. Peptide aldehydes and peptide alpha-keto esters containing a hydrophobic residue in the P1 position have been tested as potential inhibitors of the chymotrypsin-like activity. Three peptide aldehydes (benzyloxycarbonyl)-Leu-Leu-phenylalaninal (Z-LLF-CHO), N-acetyl-Leu-Leu-norleucinal (Ac-LLnL-CHO), and N-acetyl-Leu-Leu-methioninal (Ac-LLM-CHO) were found to be slow-binding reversible inhibitors with Ki values of 0.46, 5.7, and 33 microM, respectively. The simplest kinetic model for inhibition is consistent with a mechanism involving a slow and reversible association of the enzyme with the inhibitor to form a EI complex. The aldehyde inhibitors also inhibited the trypsin-like and PGPH activities of the complex albeit with much higher Ki values than those for chymotrypsin-like activity. Z-LLF-CHO, the most selective of the three aldehydes, did not inhibit the PGPH activity at concentrations of up to 200 microM and inhibited the trypsin-like activity with a Ki approximately 2 orders of magnitude higher than that for the chymotrypsin-like activity. The activity of the DCI-resistant component was not affected by Z-LLF-CHO.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York 10029.
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Vinitsky
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Michaud
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JC
Powers
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Orlowski
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000187:drug effects / Q000378:metabolism*; D004399:Dynorphins / Q000378:metabolism; D004952:Esters / Q000494:pharmacology; D006023:Glycoproteins / Q000494:pharmacology; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism; D007700:Kinetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000187:drug effects / Q000378:metabolism*; D009496:Neurotensin / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D010446:Peptide Fragments / Q000378:metabolism; D010902:Pituitary Gland / Q000201:enzymology*; D046988:Proteasome Endopeptidase Complex; D014357:Trypsin / Q000187:drug effects / Q000378:metabolism
D004952:Esters; D006023:Glycoproteins; D009097:Multienzyme Complexes; D009842:Oligopeptides; D010446:Peptide Fragments; C039304:calpain inhibitors; C077180:N-benzyloxycarbonyl-leucyl-leucyl-phenylalaninal; D007987:Gonadotropin-Releasing Hormone; D009496:Neurotensin; D004399:Dynorphins; D002918:Chymotrypsin; D014357:Trypsin; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex