Cloning, expression, purification of active recombinant human 12R-LOX enzyme and its inhibition by Acalypha indica extracts
- Published
- Accepted
- Subject Areas
- Biochemistry, Molecular Biology
- Keywords
- 12R-LOX, Acalypha indica, arachidonic acid
- Copyright
- © 2017 Fatima et al.
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2017. Cloning, expression, purification of active recombinant human 12R-LOX enzyme and its inhibition by Acalypha indica extracts. PeerJ Preprints 5:e2906v1 https://doi.org/10.7287/peerj.preprints.2906v1
Abstract
12R-LOX over-expression has been reported in various pathologies such as psoriasis, proliferative dermatitis as well as pulmonary obstructive diseases indicating that this enzyme plays significant role in pathogenesis of inflammatory diseases. 12R-LOX, therefore, is a suitable target for therapeutic intervention with potential application of its inhibitors in the treatment of skin and other inflammatory disorders. Identification of such inhibitors requires sufficient quantity of active enzyme to be produced by an easy and cost effective expression systems and further development of a robust assay system to screen inhibitors against the 12R-LOX enzyme. Therefore, in the present study, a prokaryotic expression system was developed to over-express and purify active human 12R-LOX enzyme by a single step purification process. We have further standardized an HPLC based assay system to assess the activity of purified human 12R-LOX enzyme. We show here that purified 12R-LOX preferentially utilizes free arachidonic acid as the substrate but it is also active on methyl ester of arachidonic acid, albeit less efficiently. Additionally, using this assay system we observed the potent inhibition of human 12R-LOX enzyme activity by the ethyl acetate and aqueous sub-fractions of Acalypha indica leaves, which is widely used in traditional medicines for the treatment of various skin ailments
Author Comment
This is a submission to PeerJ for review.
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