Uncoupling intramolecular processing and substrate hydrolysis in the N-terminal nucleophile hydrolase hASRGL1 by circular permutation.

Article Details

Citation

Li W, Cantor JR, Yogesha SD, Yang S, Chantranupong L, Liu JQ, Agnello G, Georgiou G, Stone EM, Zhang Y

Uncoupling intramolecular processing and substrate hydrolysis in the N-terminal nucleophile hydrolase hASRGL1 by circular permutation.

ACS Chem Biol. 2012 Nov 16;7(11):1840-7. doi: 10.1021/cb300232n. Epub 2012 Aug 29.

PubMed ID
22891768 [ View in PubMed
]
Abstract

The human asparaginase-like protein 1 (hASRGL1) catalyzes the hydrolysis of l-asparagine and isoaspartyl-dipeptides. As an N-terminal nucleophile (Ntn) hydrolase superfamily member, the active form of hASRGL1 is generated by an intramolecular cleavage step with Thr168 as the catalytic residue. However, in vitro, autoprocessing is incomplete (~50%), fettering the biophysical characterization of hASRGL1. We circumvented this obstacle by constructing a circularly permuted hASRGL1 that uncoupled the autoprocessing reaction, allowing us to kinetically and structurally characterize this enzyme and the precursor-like hASRGL1-Thr168Ala variant. Crystallographic and biochemical evidence suggest an activation mechanism where a torsional restraint on the Thr168 side chain helps drive the intramolecular processing reaction. Cleavage and formation of the active site releases the torsional restriction on Thr168, which is facilitated by a small conserved Gly-rich loop near the active site that allows the conformational changes necessary for activation.

DrugBank Data that Cites this Article

Drug Targets
DrugTargetKindOrganismPharmacological ActionActions
AsparagineIsoaspartyl peptidase/L-asparaginaseProteinHumans
No
Substrate
Details
Polypeptides
NameUniProt ID
Isoaspartyl peptidase/L-asparaginaseQ7L266Details