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Title: Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3)
Contributor(s): Alghamdi, Othman A (author); King, Nicola (author); Jones, Graham L  (author)orcid ; Moens, Pierre  (author)orcid 
Publication Date: 2017
DOI: 10.1007/s00232-017-9990-x
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Abstract: Tri- and dipeptides are transported in the kidney by PEPT1 and PEPT2 isoforms. The aim of this study was to investigate differences in transport kinetics between renal brush border (BBMV) and outer medulla (OMMV) membrane vesicles (where PEPT1 and PEPT2 are sequentially available) for a range of di- and tripeptides and peptidomimetic drugs. This was accomplished through the use of the potential-sensitive fluorescent dye 3,3'-dipropylthiacarbocyanine iodide [DiS-C₃-(3)]. BBMV and OMMV were prepared from the rat kidney using standard techniques. The presence of PEPT1 in BBMV and PEPT2 in OMMV was confirmed using Western blotting. Fluorescence changes were measured when extravesicular medium at pH 6.6 containing 0-1 mM substrates was added to a cuvette containing vesicles pre-equilibrated at pH 7.4 and 2.71 µM DiS-C₃-(3). An increase in fluorescence intensity occurred upon substrate addition reflecting the expected positive change in membrane potential difference. Of the range of substrates studied, OMMV manifested the highest affinity to cefadroxil and valacyclovir (Kₘ 4.3 ± 1.2 and 11.7 ± 3.2 µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (Kₘ 15.4 ± 3.1 µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (Kₘ 47.1 ± 9.8 µM, 55.5 ± 2.8 ΔF/s/mg protein) than BBMV (Kₘ 78.1 ± 13.3 µM and 35.5 ± 1.7 ΔF/s/mg protein, respectively). In conclusion, this study successfully separated the expression of PEPT1 and PEPT2 into different vesicle preparations inferring their activity in different regions of the renal proximal tubule.
Publication Type: Journal Article
Source of Publication: Journal of Membrane Biology, 250(6), p. 641-649
Publisher: Springer New York LLC
Place of Publication: United States of America
ISSN: 0022-2631
Field of Research (FOR): 060101 Analytical Biochemistry
110101 Medical Biochemistry: Amino Acids and Metabolites
110106 Medical Biochemistry: Proteins and Peptides (incl. Medical Proteomics)
Socio-Economic Outcome Codes: 920199 Clinical Health (Organs, Diseases and Abnormal Conditions) not elsewhere classified
Peer Reviewed: Yes
HERDC Category Description: C1 Refereed Article in a Scholarly Journal
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