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. Author manuscript; available in PMC: 2011 Aug 1.
Published in final edited form as: Mol Biosyst. 2010 Apr 14;6(8):1411–1418. doi: 10.1039/c000237b

Figure 2. GP-NAE phosphodiesterase activity is lost in GDE1(−/−) tissue.

Figure 2

(A) Near-complete loss of conversion of 14C-GP-NAE to 14C-NAE is evident from a representative thin layer radiochromatogram (left) and quantified data from replicate experiments (right). (B) Loss of conversion of 14C-lysoNAPE to 14C-NAE is evident from a representative thin layer radiochromatogram (left) and quantified data from replicate experiments (right). Note also the accumulation of the GP-NAE intermediate in the assay with GDE1(−/−) tissue. All assays were performed in PBS buffer with tissue homogenates from brains of GDE1(+/+) and (−/−) mice. (C) Ex vivo assay demonstrating that EDTA induces GP-NAE accumulation in brain homogenates due to disruption of GDE1 activity. Brain tissue from GDE1(+/−) and (−/−) mice was homogenized in the presence or absence of 10 mM EDTA (which inhibits GDE1) and incubated at room temperature for 4 hours. The addition of EDTA leads to a rapid accumulation of GP-NAEs in wild-type brain, as previously reported. GP-NAE accumulation was observed in brain tissue from GDE1(−/−) mice with- or without EDTA, demonstrating that GDE1 is the EDTA-sensitive GP-NAE phosphodiesterase in wild-type tissue. n = 4 mice per group. Results are presented as means ± standard error. Asterisks designate p < 0.05, p < 0.01, p < 0.005 (student’s t-test) for *, **, and ***, respectively. Asterisks represent difference from wild-type in panels (A) and (B), and difference from EDTA-treated GDE1(+/−) tissue in panel (C).