Inhibition of mammalian legumain by Michael acceptors and AzaAsn-halomethylketones.
Legumain is a lysosomal cysteine peptidase specific for an asparagine residue in the P1-position. It has been classified as a member of clan CD peptidases due to predicted structural similarities to caspases and gingipains. So far, inhibition studies on legumain are limited by the use of endogenous inhibitors such as cystatin C. A series of Michael acceptor inhibitors based on the backbone Cbz-L-Ala-L-Ala-L-Asn (Cbz= benzyloxycarbonyl) has been prepared and resulted in an irreversible inhibition of porcine legumain. Variation of the molecular size within the 'war head' revealed the best inhibition for the compound containing the allyl ester (kobs/I=766 M(-1) s(-1)). To overcome cyclisation between the amide moiety of the Asn residue and the 'war head', several asparagine analogues have been synthesised. Integrated in halomethylketone inhibitors, azaasparagine is accepted by legumain in the P1-position. The most potent inhibitor of this series, Cbz-L-Ala-L-Ala-AzaAsn-chloromethylketone, displays a k(obs)/I value of 139,000 M(-1) s(-1). Other cysteine peptidases, such as papain and cathepsin B, are not inhibited by this compound at concentrations up to 100 microM. The synthetic inhibitors described here represent useful tools for the investigation of the structural and physiological properties of this unique asparagine-specific peptidase.
Niestroj
André J
AJ
Probiodrug AG, Biocenter, Halle, Saale, Germany.
Feussner
Kirstin
K
Heiser
Ulrich
U
Dando
Pam M
PM
Barrett
Alan
A
Gerhartz
Bernd
B
Demuth
Hans-Ulrich
HU
D000818:Animals; D001216:Asparagine; D003546:Cysteine Endopeptidases / Q000737:chemistry*; D005456:Fluorescent Dyes; D007659:Ketones / Q000138:chemical synthesis* / Q000737:chemistry; D007700:Kinetics; D009842:Oligopeptides / Q000138:chemical synthesis* / Q000737:chemistry; D011480:Protease Inhibitors / Q000138:chemical synthesis* / Q000737:chemistry; D013329:Structure-Activity Relationship; D013552:Swine
D005456:Fluorescent Dyes; D007659:Ketones; D009842:Oligopeptides; D011480:Protease Inhibitors; D001216:Asparagine; D003546:Cysteine Endopeptidases