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Journal of Lipids
Volume 2012, Article ID 610937, 13 pages
http://dx.doi.org/10.1155/2012/610937
Research Article

Solubilization and Humanization of Paraoxonase-1

1Department of Chemistry, The Ohio State University, Columbus, OH 43210, USA
2Physiology and Immunology Branch, Research Division, United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010, USA
3Department of Biochemistry, The Ohio State University, Columbus, OH 43210, USA

Received 12 January 2012; Revised 26 March 2012; Accepted 26 March 2012

Academic Editor: Alejandro Gugliucci

Copyright © 2012 Mohosin Sarkar et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Paraoxonase-1 (PON1) is a serum protein, the activity of which is related to susceptibility to cardiovascular disease and intoxication by organophosphorus (OP) compounds. It may also be involved in innate immunity, and it is a possible lead molecule in the development of a catalytic bioscavenger of OP pesticides and nerve agents. Human PON1 expressed in E. coli is mostly found in the insoluble fraction, which motivated the engineering of soluble variants, such as G2E6, with more than 50 mutations from huPON1. We examined the effect on the solubility, activity, and stability of three sets of mutations designed to solubilize huPON1 with fewer overall changes: deletion of the N-terminal leader, polar mutations in the putative HDL binding site, and selection of the subset of residues that became more polar in going from huPON1 to G2E6. All three sets of mutations increase the solubility of huPON1; the HDL-binding mutant has the largest effect on solubility, but it also decreases the activity and stability the most. Based on the G2E6 polar mutations, we “humanized” an engineered variant of PON1 with high activity against cyclosarin (GF) and found that it was still very active against GF with much greater similarity to the human sequence.