2020
DOI: 10.3390/ijms21197094
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Sub-Nanometer Cryo-EM Density Map of the Human Heterodimeric Amino Acid Transporter 4F2hc-LAT2

Abstract: Heterodimeric amino acid transporters (HATs) are protein complexes mediating the transport of amino acids and derivatives thereof across biological membranes. HATs are composed of two subunits, a heavy and a light chain subunit belonging to the solute carrier (SLC) families SLC3 and SLC7. The human HAT 4F2hc-LAT2 is composed of the type-II membrane N-glycoprotein 4F2hc (SCL3A2) and the L-type amino acid transporter LAT2 (SLC7A8), which are covalently linked to each other by a conserved disulfide bridge. Wherea… Show more

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Cited by 10 publications
(19 citation statements)
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References 42 publications
(93 reference statements)
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“…The chromatogram of 4F2hc-LAT1 displayed a prominent, sharp peak, indicating that the double affinity purification procedure applied yielded a pure and monodisperse membrane protein sample (Figure 3A). The elution volume was similar to the GDN-solubilized heterodimeric complex of 4F2hc-LAT2, as previously reported [16]. Since 4F2hc-LAT1 and 4F2hc-LAT2 share a high sequence similarity of 48% and are structurally similar [14,16,20], these data, together with the results obtained from SDS-PAGE and Western blot analysis (Figure 2) further, corroborated correct assembly of the heterodimeric complex 4F2hc-LAT1.…”
Section: Characterization Of Purified Human 4f2hc-lat1 Proteinsupporting
confidence: 88%
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“…The chromatogram of 4F2hc-LAT1 displayed a prominent, sharp peak, indicating that the double affinity purification procedure applied yielded a pure and monodisperse membrane protein sample (Figure 3A). The elution volume was similar to the GDN-solubilized heterodimeric complex of 4F2hc-LAT2, as previously reported [16]. Since 4F2hc-LAT1 and 4F2hc-LAT2 share a high sequence similarity of 48% and are structurally similar [14,16,20], these data, together with the results obtained from SDS-PAGE and Western blot analysis (Figure 2) further, corroborated correct assembly of the heterodimeric complex 4F2hc-LAT1.…”
Section: Characterization Of Purified Human 4f2hc-lat1 Proteinsupporting
confidence: 88%
“…In addition, negligible contributions of mainly 4F2hc monomers were discerned (Figure 2, lane 1). The latter observation of minor amounts of free 4F2hc and also possibly LAT1 might reflect SDS-based denaturation of the complex due to SDS-PAGE, which was previously observed by SDS-and blue native PAGE, and corroborated by size exclusion chromatography (SEC) of 4F2hc-LAT2 [16,19]. Western blots probed with either αHis (Figure 2, lane 3) or αStrep antibodies (Figure 2, lane 5) under non-reducing conditions confirmed the high purity and homogeneity of 4F2hc-LAT1 by also displaying prominent bands at 140 kDa in either case.…”
Section: Large-scale Expression and Purificationsupporting
confidence: 75%
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“…Recent high resolution cryogenic electron microscopy (cryo-EM) structurers of L-type amino acid transporters LAT1/4F2hc, LAT2/4F2hc and b 0,+ AT/rBAT revealed the structural organisation of the heterodimer [26][27][28][29][30][31] , with the large glycosylated ectodomain sitting atop the transporter. The LAT subunits are members of the amino acid polyamine cation (APC) superfamily 32 , which exhibit a conserved '5+5 inverted topology' fold, wherein the first five TM helices are related to the second five helices via a pseudo two-fold symmetry axis running parallel to the plane of the membrane 33 .…”
Section: Introductionmentioning
confidence: 99%