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This content will become publicly available on January 1, 2026

Title: Development and use of DJ-1 affinity microcolumns to screen and study small drug candidates for Parkinson's disease
Background: DJ-1 is a protein whose mutation causes rare heritable forms of Parkinson’s disease (PD) and is of interest as a target for treating PD and other disorders. This work used high performance affinity microcolumns to screen and examine the binding of small molecules to DJ-1, as could be used to develop new therapeutics or to study the role of DJ-1 in PD. Non-covalent entrapment was used to place microgram quantities of DJ-1 in an unmodified form within microcolumns, which were then used in multiple studies to analyze binding by model compounds and possible drug candidates to DJ-1. Results: Several factors were examined in optimizing the entrapment method, including the addition of a reducing agent to maintain a reduced active site cysteine residue in DJ-1, the concentration of DJ-1 employed, and the entrapment times. Isatin was used as a known binding agent (dissociation constant, ~2.0 µM) and probe for DJ-1 activity. This compound gave good retention on 2.0 cm × 2.1 mm inner diameter DJ-1 microcolumns made under the final entrapment conditions, with a typical retention factor of 14 and elution in ~8 min at 0.50 mL/min. These DJ-1 microcolumns were used to evaluate the binding of small molecules that were selected in silico to bind or not to bind DJ-1. A compound predicted to have good binding with DJ-1 gave a retention factor of 122, an elution time of ~15 min at 0.50 mL/min, and an estimated dissociation constant for this protein of 0.5 µM. Significance: These chromatographic tools can be used in future work to screen additional possible binding agents for DJ-1 or adapted for examining drug candidates for other proteins. This work represents the first time protein entrapment has been deployed with DJ-1, and it is the first experimental confirmation of binding to DJ-1 by a small lead compound selected in silico.  more » « less
Award ID(s):
2404209 2320239 2108881
PAR ID:
10559090
Author(s) / Creator(s):
; ; ; ; ; ; ;
Publisher / Repository:
Elsevier
Date Published:
Journal Name:
Analytica Chimica Acta
Volume:
1336
Issue:
C
ISSN:
0003-2670
Page Range / eLocation ID:
343520
Subject(s) / Keyword(s):
DJ-1 Parkinson’s disease Drug screening Affinity microcolumn Protein entrapment In silico drug design
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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