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 0-1mM 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 (K m 4.3±1.2 and 11.7±3.2µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (K m 15.4±3.1µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (K m 47.1±9.8µM, 55.5±2.8ΔF/s/mg protein) than BBMV (K m 78.1±13.3µM and 35.5±1.7ΔF/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.
Original language | English |
---|---|
Pages (from-to) | 641-649 |
Number of pages | 0 |
Journal | Journal of Membrane Biology |
Volume | 250 |
Issue number | 6 |
Early online date | 7 Oct 2017 |
DOIs | |
Publication status | Published - Dec 2017 |
Keywords
- DiS-C3-(3)
- PEPT cotransporters
- Renal membrane vesicles
Fingerprint
Dive into the research topics of 'Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3).'. Together they form a unique fingerprint.
View full fingerprint
Cite this
- APA
- Author
- BIBTEX
- Harvard
- Standard
- RIS
- Vancouver
Alghamdi, O. A., King, N., Jones, G. L., & Moens, P. D. J. (2017). Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3). Journal of Membrane Biology, 250(6), 641-649. https://doi.org/10.1007/s00232-017-9990-x
Alghamdi, Othman A. ; King, Nicola ; Jones, Graham L. et al. / Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3). In: Journal of Membrane Biology. 2017 ; Vol. 250, No. 6. pp. 641-649.
@article{9ac9f638f7974521b5486e00a3c12e37,
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).",
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 0-1mM 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 (K m 4.3±1.2 and 11.7±3.2µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (K m 15.4±3.1µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (K m 47.1±9.8µM, 55.5±2.8ΔF/s/mg protein) than BBMV (K m 78.1±13.3µM and 35.5±1.7ΔF/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.",
keywords = "DiS-C3-(3), PEPT cotransporters, Renal membrane vesicles",
author = "Alghamdi, {Othman A.} and Nicola King and Jones, {Graham L.} and Moens, {Pierre D.J.}",
year = "2017",
month = dec,
doi = "10.1007/s00232-017-9990-x",
language = "English",
volume = "250",
pages = "641--649",
journal = "Journal of Membrane Biology",
issn = "0022-2631",
publisher = "Springer New York",
number = "6",
}
Alghamdi, OA, King, N, Jones, GL & Moens, PDJ 2017, 'Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3).', Journal of Membrane Biology, vol. 250, no. 6, pp. 641-649. https://doi.org/10.1007/s00232-017-9990-x
Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3). / Alghamdi, Othman A.; King, Nicola; Jones, Graham L. et al.
In: Journal of Membrane Biology, Vol. 250, No. 6, 12.2017, p. 641-649.
Research output: Contribution to journal › Article › peer-review
TY - JOUR
T1 - Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3).
AU - Alghamdi, Othman A.
AU - King, Nicola
AU - Jones, Graham L.
AU - Moens, Pierre D.J.
PY - 2017/12
Y1 - 2017/12
N2 - 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 0-1mM 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 (K m 4.3±1.2 and 11.7±3.2µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (K m 15.4±3.1µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (K m 47.1±9.8µM, 55.5±2.8ΔF/s/mg protein) than BBMV (K m 78.1±13.3µM and 35.5±1.7ΔF/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.
AB - 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 0-1mM 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 (K m 4.3±1.2 and 11.7±3.2µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (K m 15.4±3.1µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (K m 47.1±9.8µM, 55.5±2.8ΔF/s/mg protein) than BBMV (K m 78.1±13.3µM and 35.5±1.7ΔF/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.
KW - DiS-C3-(3)
KW - PEPT cotransporters
KW - Renal membrane vesicles
U2 - 10.1007/s00232-017-9990-x
DO - 10.1007/s00232-017-9990-x
M3 - Article
SN - 0022-2631
VL - 250
SP - 641
EP - 649
JO - Journal of Membrane Biology
JF - Journal of Membrane Biology
IS - 6
ER -
Alghamdi OA, King N, Jones GL, Moens PDJ. Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C3-(3). Journal of Membrane Biology. 2017 Dec;250(6):641-649. Epub 2017 Oct 7. doi: 10.1007/s00232-017-9990-x