Document Detail

Transmembrane helix 1 contributes to substrate translocation and protein stability of bile acid transporter SLC10A2.
MedLine Citation:
PMID:  21646357     Owner:  NLM     Status:  MEDLINE    
The human apical sodium-dependent bile acid transporter (hASBT, SLC10A2) plays a critical role in the enterohepatic circulation of bile acids, as well as in cholesterol homeostasis. ASBT reclaims bile acids from the distal ileum via active sodium co-transport, in a multistep process, orchestrated by key residues in exofacial loop regions, as well as in membrane-spanning helices. Here, we unravel the functional contribution of highly conserved transmembrane helix 1 (TM1) on the hASBT transport cycle. Consecutive cysteine substitution of individual residues along the TM1 helix (Ile(29)-Gly(50)), as well as exofacial Asn(27) and Asn(28), resulted in functional impairment of ∼70% of mutants, despite appreciable cell surface expression for all but G50C. Cell surface expression of G50C and G50A was rescued upon MG132 treatment as well as cyclosporine A, but not by FK506 or bile acids, suggesting that Gly(50) is involved in hASBT folding. TM1 accessibility to membrane-impermeant MTSET remains confined to the exofacial half of the helix along a single, discrete face. Substrate protection from MTSET labeling was temperature-dependent for L34C, T36C, and L38C, consistent with conformational changes playing a role in solvent accessibility for these mutants. Residue Leu(30) was shown to be critical for both bile acid and sodium affinity, while Asn(27), Leu(38), Thr(39), and Met(46) participate in sodium co-transport. Combined, our data demonstrate that TM1 plays a pivotal role in ASBT function and stability, thereby providing further insight in its dynamic transport mechanism.
Tatiana Claro da Silva; Naissan Hussainzada; Chandra M Khantwal; James E Polli; Peter W Swaan
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2011-06-06
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  286     ISSN:  1083-351X     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2011 Aug 
Date Detail:
Created Date:  2011-08-01     Completed Date:  2011-10-10     Revised Date:  2013-07-18    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  27322-32     Citation Subset:  IM    
Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland 21201, USA.
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MeSH Terms
Amino Acid Sequence
COS Cells
Cell Line
Cercopithecus aethiops
Lysosomes / metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed
Organic Anion Transporters, Sodium-Dependent / chemistry,  genetics,  metabolism*
Proteasome Endopeptidase Complex / metabolism
Protein Transport
Sequence Homology, Amino Acid
Symporters / chemistry,  genetics,  metabolism*
Grant Support
Reg. No./Substance:
0/Organic Anion Transporters, Sodium-Dependent; 0/Symporters; 145420-23-1/sodium-bile acid cotransporter; EC Endopeptidase Complex

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