Format

Send to

Choose Destination
Phytochem Anal. 2016 Mar-Apr;27(2):126-32. doi: 10.1002/pca.2607. Epub 2016 Jan 18.

A luminescence assay for natural product inhibitors of the Mycobacterium tuberculosis proteasome.

Author information

1
Department of Chemistry and Biochemistry, University of the Sciences, Philadelphia, PA, USA.
2
Department of Biological Sciences, University of the Sciences, Philadelphia, PA, USA.

Abstract

INTRODUCTION:

Mycobacterium tuberculosis (Mtb) causes a large global burden of disease, with a high mortality rate in healthy and immuno-compromised patients. A number of molecular targets have been identified for treatment of this disease, including the Mtb proteasome. The Mtb proteasome enhances Mtb survival during nitrosative and oxidative stress in the latent, non-replicative phase. Therefore, Mtb proteasome inhibition could help to combat Mtb infections that do not respond to current therapies.

OBJECTIVE:

To develop and validate a novel biochemical assay to assess Mtb proteasome activity in the presence of organic and aqueous plant test extracts.

METHOD:

Fluorescence (photoluminescence) and luminescence (chemiluminescence) assays were investigated as potential methods to determine the robustness and repeatability for use in screening natural product extracts for Mtb proteasome inhibitors.

RESULTS:

The fluorescence assay, used widely for Mtb proteasome activity assays, was subject to interference due to the natural fluorescence of compounds in many of the extracts; there is little interference with the luminescence approach. As proof of principle, we used the luminescence assay to screen a small set of plant test extracts.

CONCLUSIONS:

Luminescence is the more suitable assay for assay of plant natural product extracts. The sensitivities of the luminescence and fluorescence assays are comparable. A Z'-factor of 0.58 for the luminescence assay makes it suitable for medium-to-high throughput screening efforts.

KEYWORDS:

Mycobacterium tuberculosis; luminescence; natural products; proteasome

PMID:
26778282
DOI:
10.1002/pca.2607
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center