Human Paraoxonase-3 Is an HDL-Associated Enzyme With Biological Activity Similar to Paraoxonase-1 Protein but Is Not Regulated by Oxidized Lipids

Reddy, Srinivasa T.; Wadleigh, David J.; Grijalva, Víctor; Ng, Carey J.; Hama, Susan; Gangopadhyay, Aditya; Shih, Diana M.; Lusis, Aldons J.; Navab, Mohamad; Fogelman, Alan M.

doi:10.1161/01.atv.21.4.542

Cited by 315 publications

(297 citation statements)

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“…Activity was very low (Ͻ0.1% of that of PON1) and exhibited a prolonged lag (Ϸ30 min) in onset (hysteresis). These two factors may account for the earlier reports of no paraoxonase activity in the serum-purified RabPON3 (13,14). Thus, the enzymatic properties of RabPON3 are not significantly altered on fusion to thioredoxin and expression in E. coli, and the detailed kinetic parameters obtained here are, in general, relevant to wild-type PON3s.…”

Section: Newly Evolved Pon3 Variants (Repon3)supporting

confidence: 69%

“…Plasmids containing the HuPON1 and HuPON3 genes (14) were used as templates for PCR amplification. The genes for MoPON1, MoPON3, and RatPON1 were amplified from mouse and rat liver cDNA (Clontech).…”

Section: Methodsmentioning

confidence: 99%

“…Wild-type PON3s exhibit 100-to 1,000-fold lower rates of ester and phosphotriester hydrolysis than PON1, although lactonase activities of the two enzymes are similar (13,14). Their sequences are Ϸ65% identical at the amino acid level.…”

Section: Newly Evolved Pon3 Variants (Repon3)mentioning

confidence: 99%

“…Because of its low level in the serum (Ϸ50-fold lower than PON1), PON3 was only recently purified and characterized (13,14), and it was not expressed in heterologous expression systems. The specific kinetic parameters of serum PON3 have not been determined, but relative activities of various substrates of RabPON3 are available (13).…”

Section: Newly Evolved Pon3 Variants (Repon3)mentioning

confidence: 99%

“…PON3's low level in the high-density lipoprotein (Ϸ50-fold lower than PON1) hinders its purification and detailed characterization. PON3s were only recently purified and characterized from human and rabbit sera (13,14). They were found to hydrolyze a wide variety of esters and lactones but not paraoxon.…”

mentioning

confidence: 99%

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Directed evolution of mammalian paraoxonases PON1 and PON3 for bacterial expression and catalytic specialization

Aharoni

Gaidukov

Yagur

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

266

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Serum paraoxonases (PONs) are a group of enzymes that play a key role in organophosphate (OP) detoxification and in prevention of atherosclerosis. However, their structure and mechanism of action are poorly understood. PONs seem like jacks-of-all-trades, acting on a very wide range of substrates, most of which are of no physiological relevance. Family shuffling and screening lead to the first PON variants that express in a soluble and active form in Escherichia coli. We describe variants with kinetic parameters similar to those reported for PONs purified from sera and others that show dramatically increased activities. In particular, we have evolved PON1 variants with OP-hydrolyzing activities 40-fold higher than wild type and a specificity switch of >2,000-fold, producing PONs specialized for OP rather than ester hydrolysis. Analysis of the newly evolved variants provides insights into the evolutionary relationships between different family members.

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Section: Newly Evolved Pon3 Variants (Repon3)supporting

confidence: 69%

Section: Methodsmentioning

confidence: 99%

Section: Newly Evolved Pon3 Variants (Repon3)mentioning

confidence: 99%