Asparagine endopeptidase can initiate the removal of the MHC class II invariant chain chaperone.

The invariant chain (Ii) chaperone for MHC class II molecules is crucial for their effective function. Equally important is its removal. Cathepsins S or L are known to be required for the final stages of Ii removal in different APCs, but the enzymes which initiate Ii processing have not been identified. Here we show that this step can be performed in B lymphocytes by asparagine endopeptidase (AEP), which targets different asparagine residues in the lumenal domain of human and mouse invariant chain. Inhibition of AEP activity slows invariant chain processing and hinders the expression of an antigenic peptide engineered to replace the groove binding region of Ii (CLIP). However, the initiation of Ii removal can also be performed by other proteases, reflecting the importance of this step.

Novel cell-permeable acyloxymethylketone inhibitors of asparaginyl endopeptidase.

Mammalian asparaginyl endopeptidase (AEP) or legumain is a recently identified lysosomal cysteine protease belonging to clan CD. To date it has been shown to be involved in antigen presentation within class II MHC positive cells and in pro-protein processing. Further elucidation of the biological functions of the enzyme will require potent and selective inhibitors and thus we describe here new acyloxymethylketone inhibitors of AEP. The most potent of the series is 2,6-dimethyl-benzoic acid 3-benzyloxycarbonylamino-4-carbamoyl-2-oxo-butyl ester (MV026630) with a kobs/[I] value of 1.09 x 10(5) M(-1) s(-1). At low microM concentrations this compound is able to enter living cells and irreversibly inactivate AEP. We show that this results in inhibition of AEP autoactivation and in perturbation of the processing and presentation of T cell epitopes from both tetanus toxin and myelin basic protein.