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Molecular cloning and expression of the C-terminal domain of mouse NTE-related esterase

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Abstract

NTE-related esterase (NRE), conserved in mouse, rat and human, was a member of patatin-like phospholipases (PLPLA) with high homology to neuropathy target esterase (NTE). Little has been known about the characteristics of NRE and NRE functional esterase activity has yet not been defined. The C-terminal gene sequence of mouse NRE (mNREC) encoding 923–1,326 amino acid containing the patatin domain was first cloned and then expressed tagged with enhanced green fluorescence protein (EGFP) in mammalian cells. The results showed that mNREC had NTE esterase activity in mammalian cells. Overexpression of mNREC did not affect the esterase activity sensitive to paraoxon or resistant to both paraoxon and mipafox. mNREC was distributed in the cytoplasm in contrast to the distribution of human NTE esterase domain. The expression analysis of NRE gene in adult mouse tissues by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) showed that there were higher levels of NRE mRNA in the brain and testis than in the liver and kidney, which was about 50% and 35% of that in the brain. These results firstly showed the tissue distribution of NRE gene in adult mouse and defined that NRE had functional esterase activity.

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Abbreviations

DMEM:

Dulbecco’s modified Eagle’s medium

EDTA:

Ethylenediaminetetraacetic acid

ECL:

Enhanced chemiluminescence

EGFP:

Enhanced green fluorescence protein

ER:

Endoplasmic reticulum

GPC:

Glycerophosphocholine

NEST:

NTE esterase activity domain

NP40:

Nonidet P-40

NTE:

Neuropathy target esterase

NRE:

NTE-related esterase

NREC:

C-terminal domain of NRE

OPIDN:

OP-induced delayed neuropathy

PBS:

Phosphate-buffered saline

PC:

Phosphatidylcholine

PCR:

Polymerase chain reaction

PLPLA:

Patatin-like phospholipases

PV:

Phenyl valerate

RT-PCR:

Reverse transcription-PCR

SDS-PAGE:

Sodium dodecyl sulphate-polyacrylamide gel electrophoresis

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Acknowledgments

This work was supported by grants from the National Nature Science Foundation of China (30600329) and the Natural Science Foundation of Chongqing City (CSTC2005BB5072) and the Science and Technology Project from Chongqing Municipal Education committee (KJ070510). The authors would like to thank Dr. Paul Glynn for kindly providing D16 plasmid of human NTE cDNA.

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Authors and Affiliations

  1. Key Laboratory of Molecular Biology, College of Bio-information, Chongqing University of Posts and Telecommunications, Chongqing, 400065, P.R. China

    Ping-An Chang

  2. Laboratory of Molecular Toxicology, Institute of Zoology, Chinese Academy of Sciences, Beijing, 100080, P.R. China

    Ping-An Chang, Ding-Xin Long & Yi-Jun Wu

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  1. Ping-An Chang
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  2. Ding-Xin Long
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Correspondence to Ping-An Chang.

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Chang, PA., Long, DX. & Wu, YJ. Molecular cloning and expression of the C-terminal domain of mouse NTE-related esterase. Mol Cell Biochem 306, 25–32 (2007). https://doi.org/10.1007/s11010-007-9550-2

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  • DOI: https://doi.org/10.1007/s11010-007-9550-2

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