Mutagenesis and cysteine scanning of transmembrane domain 10 of the human dipeptide transporter.
Mutagenesis and cysteine scanning of transmembrane domain 10 of the human dipeptide transporter.
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DOI：
10.1007/s11095-009-9952-9
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发表时间：
2009-10
影响因子：
3.7
通讯作者：
Davies, Daryl L.
中科院分区：
文献类型：
--
作者：
Xu, Liya;Haworth, Ian S.;Kulkarni, Ashutosh A.;Bolger, Michael B.;Davies, Daryl L.
关键词：
The human dipeptide transporter (hPEPT1) facilitates transport of dipeptides and drugs from the intestine into the circulation. The role of transmembrane domain 10 (TMD10) of hPEPT1 in substrate translocation was investigated using cysteine-scanning mutagenesis with 2-Trimethylammonioethyl methanethiosulfonate (MTSET). Each amino acid in TMD10 was mutated individually to cysteine, and transport of [3H]Gly-Sar was evaluated with and without MTSET following transfection of each mutant in HEK293 cells. Similar localization and expression levels of wild type (WT) hPEPT1 and all mutants were confirmed by immunostaining and biotinylation followed by western blot analysis. E595C- and G594C-hPEPT1 showed negligible Gly-Sar uptake. E595D-hPEPT1 showed similar uptake to WT-hPEPT1, but E595K- and E595R-hPEPT1 did not transport Gly-Sar. Double mutations E595K/R282E and E595R/R282E did not restore uptake. G594A-hPEPT1 showed similar uptake to WT-hPEPT1, but G594V-hPEPT1 eliminated uptake. Y588C-hPEPT1 showed uptake of 20% that of WT-hPEPT1. MTSET modification supported a model of TMD10 with an amphipathic helix from I585 to V600 and increased solvent accessibility from T601 to F605. Our results suggest that G594 and E595 in TMD10 of hPEPT1 have key roles in substrate transport and that Y588 may have an important secondary mechanistic role.
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影响因子：
4.8
作者：
RONG, LA;FEI, YJ;LEIBACH, FH
通讯作者：
LEIBACH, FH
影响因子：
4.5
作者：
Links, Jennifer L. S.;Kulkarni, Ashutosh A.;Haworth, Ian S.
通讯作者：
Haworth, Ian S.
精氨酸282的位置定向诱变表明质子和肽是如何通过兔Pept1共同传输的。
DOI：
10.1016/j.biocel.2007.10.010
发表时间：
2008
影响因子：
4
作者：
Pieri, Myrtani;Hall, Dashiell;Price, Richard;Bailey, Patrick;Meredith, David
通讯作者：
Meredith, David
DOI：
10.1016/s0006-291x(03)00926-4
发表时间：
2003-06-20
影响因子：
3.1
作者：
Kulkarni, AA;Haworth, IS;Lee, VHL
通讯作者：
Lee, VHL
影响因子：
2.9
作者：
Covitz, KMY;Amidon, GL;Sadée, W
通讯作者：
Sadée, W