Structural model for the organization of the transmembrane spans of the human red-cell anion exchanger (band 3; AE1)

Groves, Jonathan D.; Tanner, Michael

doi:10.1042/bj3440699

Cited by 27 publications

(24 citation statements)

References 28 publications

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“…6). Previous studies have suggested that band 3 contains subdomains comprising TM1-5, TM6 -8, and TM9 -12 that interact with each other (33)(34)(35). The presence of these extensive interactions has been confirmed by studies demonstrating that the SAO deletion causes structural perturbations throughout band 3 that were particularly evident in the C-terminal portion of the molecule (20).…”

Section: A Cytoplasmic Surface Region On the N-terminal Side Of The Msupporting

confidence: 52%

The N-terminal Region of the Transmembrane Domain of Human Erythrocyte Band 3

Kanki¹,

Young

Sakaguchi

et al. 2003

Journal of Biological Chemistry

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We studied the role of the N-terminal region of the transmembrane domain of the human erythrocyte anion exchanger (band 3; residues 361-408) in the insertion, folding, and assembly of the first transmembrane span (TM1) to give rise to a transport-active molecule. We focused on the sequence around the 9-amino acid region deleted in Southeast Asian ovalocytosis (Ala-400 to Ala-408), which gives rise to nonfunctional band 3, and also on the portion of the protein N-terminal to the transmembrane domain (amino acids 361-396). We examined the effects of mutations in these regions on endoplasmic reticulum insertion (using cell-free translation), chloride transport, and cell-surface movement in Xenopus oocytes. We found that the hydrophobic length of TM1 was critical for membrane insertion and that formation of a transport-active structure also depended on the presence of specific amino acid sequences in TM1. Deletions of 2 or 3 amino acids including Pro-403 retained transport activity provided that a polar residue was located 2 or 3 amino acids on the C-terminal side of Asp-399. Finally, deletion of the cytoplasmic surface sequence G 381 LVRD abolished chloride transport, but not surface expression, indicating that this sequence makes an essential structural contribution to the anion transport site of band 3.The red cell anion exchanger (band 3, AE1) is composed of two distinct domains: a C-terminal transmembrane domain predicted to span the membrane up to 14 times and a Nterminal cytosolic domain (1-4). The transmembrane domain mediates chloride-bicarbonate exchange, whereas the cytosolic domain associates peripheral proteins with the membrane.Southeast Asian ovalocytosis (SAO) 1 is caused by the heterozygous presence of an abnormal band 3 (band 3 SAO), which has a deletion of 9 amino acid residues (amino acids 400 -408) at the boundary between the cytosolic domain and the first transmembrane-spanning segment (5-7). SAO is prevalent in areas where malaria is endemic (8) and is thought to confer some protection against cerebral malaria (9). Homozygosity for band 3 SAO is probably lethal (10).Band 3 SAO is expressed at the surface of SAO red cells, but the mutant protein has no anion transport function and does not bind the anion transport inhibitor di-isothiocyanatodihydrostilbene either reversibly or irreversibly (11-13). Changes in di-isothiocyanatodihydrostilbene binding (14), N-glycosylation (15), and blood group antigen expression (16,17) indicate that the SAO deletion results in the misfolding of the transmembrane domain. The ␣-helical content of band 3 SAO is similar to normal band 3 (15, 18), but the transmembrane helices of band 3 SAO are disorganized while remaining folded (18).Transmembrane segment 1 (TM1) of SAO band 3 does not insert into microsomes in an alkali-stable fashion, unlike TM1 of normal band 3 (19). However, TMs 1-3 and larger fragments of SAO band 3 are incorporated stably into microsomal membranes. Lys-430, which is located in the first extracellular loop of band 3 between TM1 and TM2, is l...

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Section: A Cytoplasmic Surface Region On the N-terminal Side Of The Msupporting

confidence: 52%

The N-terminal Region of the Transmembrane Domain of Human Erythrocyte Band 3

Kanki¹,

Young

Sakaguchi

et al. 2003

Journal of Biological Chemistry

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“…Furthermore, by combining our results with co-immunoprecipitation studies indicating that TM6 is not located at the dimeric interface [29], we may deduce that TM6 is next to TMs that are at the dimeric interface.…”

Section: Discussionmentioning

confidence: 92%

“…Subunit interactions have been shown to play a role in both anion transport and inhibition of this transport by ligands [15][16][17][18]. Regions of the protein contributing to the dimeric interface have not previously been identified, although coimmunoprecipitation studies seem to exclude TMs 6-8, 13 and 14 [29]. The strategy chosen in the present study was to introduce single cysteines into putative loop regions of the protein.…”

Section: Discussionmentioning

confidence: 99%

Cysteine-directed cross-linking localizes regions of the human erythrocyte anion-exchange protein (AE1) relative to the dimeric interface

Taylor

Zhu

Casey

2001

Biochemical Journal

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The human erythrocyte anion-exchanger isoform 1 (AE1) is a dimeric membrane protein that exchanges chloride for bicarbonate across the erythrocyte plasma membrane. Crystallographic studies suggest that the transmembrane anion channel lies at the interface between the two monomers, whereas kinetic analysis provides evidence that each monomer contains an anion channel. We have studied the structure-function relationship of residues at the dimeric interface of AE1 by cysteine-directed cross-linking. Single cysteine mutations were introduced in 16 positions of putative loop regions throughout AE1. The ability of these residues to be chemically cross-linked to their partner within the dimeric protein complex was assessed by mobility of the protein on immunoblots. Introduced cysteine residues in extracellular loops (ECs) 1-4 and intracellular loop 1 formed

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“…At the molecular level, some attempts have been done to localize the part of the molecule that is involved in the formation of these complexes. Taylor et al as well as Groves et al localized this interaction region in the first four TM of hAE1 although the second one also describes other important regions (Groves and Tanner, 1999;Taylor et al, 2001). By analogy, in our model of tAE1, C462 might be involved in such a process, since it is located in the third TM.…”

Section: Discussionmentioning

confidence: 81%

Importance of several cysteine residues for the chloride conductance of trout anion exchanger 1 (tAE1)

Martial¹,

Guizouarn²,

Gabillat³

et al. 2007

Journal Cellular Physiology

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In this study, we devised a cysteine-focused point mutation analysis of the chloride channel function of trout anion exchanger 1 (tAE1) expressed in X. laevis oocytes. Seven cysteines, belonging to the transmembrane domain of tAE1, were mutated into serines (either individually or in groups) and the effects of these mutations on the chloride conductance of injected oocytes were measured. We showed that three cysteines were essential for the functional expression of tAE1. Namely, mutations C462S, C583S and C588S reduced Cl(-) conductance by 68%, 52% and 83%, respectively, when compared to wild type tAE1. These residual conductances were still inhibited by 0.5 mM niflumic acid. Western blot experiments demonstrated that C462 was involved in protein expression onto the plasma membrane. A mutant devoid of this residue was unable to express onto the plasma membrane, especially if several other cysteines were missing: consequently, the cysteine-less mutant of tAE1 was not functional. C583 and C588 were involved in the channel function of tAE1 as shown by anion substitution experiments proving that selectivity of the mutated pore differs from the wild type one. On the contrary, they were not involved in the Cl(-)/HCO(3)(-) exchange function of tAE1, as demonstrated by intracellular pH measurements. These and several complementary mutations allow us to conclude that a mutant of tAE1 containing the sole C462 can drive a marginal Cl(-) current; however, the minimal configuration necessary to get optimal functional expression of the tAE1 chloride channel is that of a mutant containing unaffected residues C462, C583 and C588.

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Structural model for the organization of the transmembrane spans of the human red-cell anion exchanger (band 3; AE1)

Cited by 27 publications

References 28 publications

The N-terminal Region of the Transmembrane Domain of Human Erythrocyte Band 3

The N-terminal Region of the Transmembrane Domain of Human Erythrocyte Band 3

Cysteine-directed cross-linking localizes regions of the human erythrocyte anion-exchange protein (AE1) relative to the dimeric interface

Importance of several cysteine residues for the chloride conductance of trout anion exchanger 1 (tAE1)

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