Transmembrane segment 5 of the dipeptide transporter hPepT1 forms a part of the substrate translocation pathway

Biochemical and Biophysical Research Communications
Ashutosh A KulkarniVincent H L Lee

Abstract

This study is the first systematic attempt to investigate the role of transmembrane segment 5 of hPepT1, the most conserved segment across different species, in forming a part of the aqueous substrate translocation pathway. We used cysteine-scanning mutagenesis in conjunction with the sulfhydryl-specific reagents, MTSEA and MTSET. Neither of these reagents reduced wild-type-hPepT1 transport activity in HEK293 cells and Xenopus oocytes. Twenty-one single cysteine mutations in hPepT1 were created by replacing each residue within TMS5 with a cysteine. HEK293 cells were then transfected with each mutated protein and the steady-state protein level, [3H]Gly-Sar uptake activity, and sensitivity to the MTS reagents were measured. S164C-, L168C-, G173C-, and I179C-hPepT1 were not expressed on the plasma membrane. Y167C-, N171C-, and S174C-hPepT1 showed </=25% Gly-Sar uptake when compared with WT-hPepT1. P182C-hPepT1 showed approximately 40% specific activity whereas all the remaining transporters, although still sensitive to single cysteine mutations, exhibited more than 50% specific activity when compared to WT-hPepT1. The activity of F166C-, L176C-, S177C-, T178C-, I180C-, T181C-, and P182C-hPepT1 was partially inhibited, while the ac...Continue Reading

References

Jun 20, 1995·Proceedings of the National Academy of Sciences of the United States of America·R T Yan, P C Maloney
Mar 1, 1995·British Journal of Pharmacology·D T ThwaitesN L Simmons
Oct 25, 1994·Proceedings of the National Academy of Sciences of the United States of America·J A JavitchA Karlin
Jun 1, 1993·Protein Science : a Publication of the Protein Society·M Sahin-Tóth, H R Kaback
Jan 9, 1996·Proceedings of the National Academy of Sciences of the United States of America·M T Pérez-GarcíaG F Tomaselli
Dec 1, 1995·British Journal of Pharmacology·U WenzelH Daniel
Mar 21, 1997·The Journal of Biological Chemistry·S NussbergerM A Hediger
Feb 7, 1998·The Journal of Biological Chemistry·C S TempleJ R Bronk
Jun 4, 1998·Biochemical and Biophysical Research Communications·M E GanapathyF H Leibach
Oct 16, 1999·Journal of Controlled Release : Official Journal of the Controlled Release Society·V H LeeC T Okamoto

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Citations

Feb 9, 2006·Pflügers Archiv : European journal of physiology·Konstantinos-E PanitsasDavid Meredith
May 12, 2011·The Journal of Biological Chemistry·Elisabeth MallePeggy C Stolt-Bergner
Mar 20, 2013·Molecular Aspects of Medicine·David E SmithMatthias A Hediger
Mar 14, 2008·Alcoholism, Clinical and Experimental Research·Kaixun LiDaryl L Davies
Feb 3, 2011·Medicinal Research Reviews·Pedro Cano-Soldado, Marçal Pastor-Anglada
Sep 28, 2014·The International Journal of Biochemistry & Cell Biology·Johanne M JensenOsman Mirza
Apr 10, 2007·The Journal of Membrane Biology·D Meredith, R A Price
Apr 18, 2008·The Journal of Pharmacy and Pharmacology·Matthias BrandschEva Bosse-Doenecke
Apr 13, 2004·The Journal of Pharmacology and Experimental Therapeutics·Eric Y ZhangKathleen M Hillgren
Mar 25, 2017·American Journal of Physiology. Gastrointestinal and Liver Physiology·Tamara StelzlHannelore Daniel
Oct 9, 2003·The Journal of Biological Chemistry·Ashutosh A KulkarniVincent H L Lee
Nov 6, 2007·The Journal of Experimental Biology·Ivar RønnestadM Leonor Cancela

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