Uncoupling intramolecular processing and substrate hydrolysis in the N-terminal nucleophile hydrolase hASRGL1 by circular permutation.
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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
Drug Target Kind Organism Pharmacological Action Actions Asparagine Isoaspartyl peptidase/L-asparaginase Protein Humans NoSubstrateDetails - Polypeptides
Name UniProt ID Isoaspartyl peptidase/L-asparaginase Q7L266 Details