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Detection and quantitation of glutamate carboxypeptidase II in human blood

Authors

  • Tomáš Knedlík,

    1. Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic
    2. Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic
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  • Václav Navrátil,

    1. Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic
    2. Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic
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  • Viktor Vik,

    1. Department of Urology, Thomayer Hospital in Prague, Prague, Czech Republic
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  • Dalibor Pacík,

    1. Department of Urology, Medical School Masaryk University, University Hospital Brno, Brno, Czech Republic
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  • Pavel Šácha,

    1. Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic
    2. Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic
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  • Jan Konvalinka

    Corresponding author
    1. Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic
    2. Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic
    • Correspondence to: Jan Konvalinka, Institute of Organic Chemistry and Biochemistry, ASCR, v.v.i. Flemingovo n. 2, Prague 6, 166 10, Czech Republic. E-mail: jan.konvalinka@uochb.cas.cz

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Abstract

BACKGROUND

Glutamate carboxypeptidase II (GCPII) is a transmembrane enzyme that cleaves N-acetyl-L-aspartyl-L-glutamate (NAAG) in the brain. GCPII is highly expressed in the prostate and prostate cancer and might be associated with prostate cancer progression. Another exopeptidase, plasma glutamate carboxypeptidase (PGCP), was reported to be similar to GCPII and to share its NAAG-hydrolyzing activity.

METHODS

We performed a radioenzymatic assay with [3H]NAAG as a substrate to detect and quantify the enzymatic activity of GCPII in plasma. Using a specific antibody raised against native GCPII (2G7), we immunoprecipitated GCPII from human plasma. We also cloned two PGCP constructs, expressed them in insect cells, and tested them for their NAAG-hydrolyzing activity.

RESULTS

We detected GCPII protein in human plasma and found that its concentration ranges between 1.3 and 17.2 ng/ml in volunteers not diagnosed with prostate cancer. Recombinant PGCP was enzymatically active but exhibited no NAAG-hydrolyzing activity.

CONCLUSION

GCPII is present in human blood, and its concentration within a healthy population varies. Recombinant PGCP does not hydrolyze NAAG, suggesting that GCPII alone is responsible for the NAAG-hydrolyzing activity observed in human blood. The potential correlation between GCPII serum levels and the disease status of prostate cancer patients will be further investigated. Prostate 74:768–780, 2014. © 2014 Wiley Periodicals, Inc.

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