Human PON1, a biomarker of risk of disease and exposure

Abstract

Human paraoxonase 1 (PON1) is a high-density lipoprotein (HDL)-associated serum enzyme that exhibits a broad substrate specificity. In addition to protecting against exposure to some organophosphorus (OP) pesticides by hydrolyzing their toxic oxon metabolites, PON1 is important in protecting against vascular disease by metabolizing oxidized lipids. Recently, PON1 has also been shown to play a role in inactivating the quorum sensing factor N-(3-oxododecanoyl)-l-homoserine lactone (3OC12-HSL) of Pseudomonas aeruginosa. Native, untagged engineered recombinant human PON1 (rHuPON1) expressed in Escherichia coli and purified by conventional column chromatographic purification is stable, active, and capable of protecting PON1 knockout mice (PON1(-/-)) from exposure to high levels of the OP compound diazoxon. The bacterially derived rHuPON1 can be produced in large quantities and lacks the glycosylation of eukaryotic systems that can produce immunogenic complications when inappropriately glycosylated recombinant proteins are used as therapeutics. Previous studies have shown that the determination of PON1 status, which reveals both PON1(192) functional genotype and serum enzyme activity level, is required for a meaningful evaluation of PON1's role in risk of disease or exposure. We have developed a new two-substrate assay/analysis protocol that provides PON1 status without use of toxic OP substrates, allowing for use of this protocol in non-specialized laboratories. Factors were also determined for inter-converting rates of hydrolysis of different substrates. PON1 status also plays an important role in revealing changes in HDL-associated PON1 activities in male patients with Parkinson disease (PD). Immunolocalization studies of PONs 1, 2 and 3 in nearly all mouse tissues suggest that the functions of PONs 1 and 3 extend beyond the plasma and the HDL particle.

Figure 1

Determination of the functional genomics of plasma PON1 using citrate stored plasma. Plotting the rates of hydrolysis of diazoxon vs. paraoxon for plasma (or serum) samples from a population divides the population into 3 distinct groups, individuals functionally homozygous for PON1_Q192, heterozygotes (PON1_Q/R192) and individuals homozygous for PON1_R192. While PCR analysis of the position 192 polymorphism may reveal that an individual possesses one copy of each allele, this analysis will pick up alleles in heterozygotes that are inactivated by any number of mutations, i.e., the analysis provides the functional status of an individual’s PON1 genomics. Reproduced from [77] with permission.

Figure 2

The human PON1 gene with known polymorphisms and their frequencies. The 5’ end of the gene is on the left. (SeattleSNPs, http://pga.gs.washington.edu/)