Die pH-Abhängigkeit des NaDC3-Transporters

pH dependency of the NaDC3 transporter

by Astrid Sophie Hoffmann
Doctoral thesis
Date of Examination:2017-12-04
Date of issue:2017-11-27
Advisor:Prof. Dr. Birgitta-Christina Burckhardt
Referee:Prof. Dr. Birgitta-Christina Burckhardt
Referee:Prof. Dr. Mladen Tzvetkov
Persistent Address: http://dx.doi.org/10.53846/goediss-6607

 

 

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Abstract

English

The sodium-dependent dicarboxylate transporter 3 (NaDC3) is located close to organic anion transporters (OAT) in the basolateral membrane of renal proximal tubule cells. The transport-activity of OAT1 (p-aminohippurate) and OAT3 (estrone-3-sulfate) is influenced by the extracellular pH.  In this study the influence of extracellular pH on substrate-induced currents of NaDC3 was investigated. The two-electrode voltage clamp technique (TEVC) was used for the experiments where substrate-induced currents in NaDC3-expressing oocytes from Xenopus laevis were measured. The results reveal that there is no effect of pH with the model substrates succinate and citrate in the range of pH 5,4-8,4. Low and high pH were less tolerated from the oocytes itselves. Michaelis-Menten constants (Km) for succinate and citrate (pH 6,5 and 7,5) were determined as well. It was shown, that there is no significant difference between pH 6,5 and 7,5, whereby NaDC3 has a higher affinity to succinate than to citrate. In a second step the study investigated the influence of pH on sodium-dependent currents. Km for sodium (pH 6,5 and 7,5) were determined and sodium was replaced by N-methyl-D-glucamine (NMDG) or choline. There was no significant difference between the Km for sodium whether sodium was substituted for NMDG or choline, but in the presence of sodium the substrate-induced currents were higher than without sodium. Then the influence of the intracellular pH due to the protonophore carbonylcynid-3-hydrazone (CCCP) was analyzed. Here the results show that there is no influence of the intracellular pH, but substrate-induced currents were lower than before after perfusion with CCCP.
Keywords: NaDC3; pH; Xenopus laevis; TEVC; SLC13A3
 
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