Aes - Explore the Science & Experts | ideXlab

Scan Science and Technology

Contact Leading Edge Experts & Companies

Aes

The Experts below are selected from a list of 297 Experts worldwide ranked by ideXlab platform

Aes – Free Register to Access Experts & Abstracts

Seth L Alper – One of the best experts on this subject based on the ideXlab platform.

  • Molecular physiology and genetics of Na+-independent SLC4 anion exchangers.
    Journal of Experimental Biology, 2009
    Co-Authors: Seth L Alper
    Abstract:

    Plasmalemmal Cl–/HCO3– exchangers are encoded by the SLC4 and SLC26 gene superfamilies, and function to regulate intracellular pH, [Cl–] and cell volume. The Cl–/HCO3– exchangers of polarized epithelial cells also contribute to transepithelial secretion and reabsorption of acid–base equivalents and Cl–. This review focuses on Na+-independent electroneutral Cl–/HCO3– exchangers of the SLC4 family. Human SLC4A1/AE1 mutations cause the familial erythroid disorders of spherocytic anemia, stomatocytic anemia and ovalocytosis. A largely discrete set of AE1 mutations causes familial distal renal tubular acidosis. The Slc4a2/Ae2–/– mouse dies before weaning with achlorhydria and osteopetrosis. A hypomorphic Ae2–/– mouse survives to exhibit male infertility with defective spermatogenesis and a syndrome resembling primary biliary cirrhosis. A human SLC4A3/AE3 polymorphism is associated with seizure disorder, and the Ae3–/– mouse has increased seizure susceptibility. The transport mechanism of mammalian SLC4/AE polypeptides is that of electroneutral Cl–/anion exchange, but trout erythroid Ae1 also mediates Cl– conductance. Erythroid Ae1 may mediate the DIDS-sensitive Cl– conductance of mammalian erythrocytes, and, with a single missense mutamutation, can mediate electrogenic SO42–/Cl– exchange. AE1 trafficking in polarized cells is regulated by phosphorylation and by interaction with other proteins. AE2 exhibits isoform-specific patterns of acute inhibition by acidic intracellular pH and independently by acidic extracellular pH. In contrast, AE2 is activated by hypertonicity and, in a pH-independent manner, by ammonium and by hypertonicity. A growing body of structure–function and interaction data, together with emerging information about physiological function and structure, is advancing our understanding of SLC4 anion exchangers.

  • Putative re-entrant loop 1 of AE2 transmembrane domain has a major role in acute regulation of anion exchange by pH.
    The Journal of biological chemistry, 2008
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Seth L Alper
    Abstract:

    Normal pH sensitivity of the SLC4A2/AE2 anion exchexchanger requires transmembrane domain (TMD) amino acid (aa) residues not conserved in the homologous but relatively pH-insensitive SLC4A1/AE1 polypeptide. We tested the hypothesis that the nonconserved aa cluster 1075DKPK1078 within the first putative re-entrant loop (RL1) of AE2 TMD contributes to pH sensor function by studying anion exchange function of AE2 mutants in which these and other RL1 aa were systematically substituted with corresponding RL1 aa from AE1. Regulation of Cl-/Cl- and Cl-/HCO(-)3 exchange by intracellular pH (pHi) or extracellular pH (pHo) was measured as 4,4′-di-isothiocyanatostilbene-2,2′ disulfonic acid-sensitive 36Cl- efflux from Xenopus oocytes. AE2 RL1 mutants 1075AAAQ1078 and 1075AAAQN1079 showed reduced pHi sensitivity and pHo sensitivity was acid-shifted by approximately 1 pH unit. Individual mutants D1075A and P1077A exhibited moderately altered pH sensitivity, whereas a range of substitutions at conserved AE2 Ile-1079 substantially altered sensitivity to pHo and/or pHi. Substitution of the complete AE1 RL1 with AE2 RL1 failed to confer AE2-like pH sensitivity onto AE1. Replacement, however, of AE1 RL1 763SGPGAAAQ770 with AE2 1071VAPGDKPK1078 restored pHi sensitivity to the chimera AE2(1-920)/AE1(613-929) without affecting its low sensitivity to pHo. The results show that acute regulation of AE2 by pH requires RL1 of the TMD. We propose that critical segments of RL1 constitute part of an AE2 pH sensor that, together with residues within the N-terminal half of the TMD, constrain the AE2 polypeptide in a conformation required for regulation of anion exchange by pHi.

  • Acute regulation of mouse AE2 anion exchanger requires isoform-specific amino acid residues from most of the transmembrane domain
    The Journal of Physiology, 2007
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Boris E. Shmukler, Seth L Alper
    Abstract:

    The widely expressed anion exchexchanger polypeptide AE2/SLC4A2 is acutely inhibited by acidic intracellular (pHi), by acidic extracellular pH (pHo), and by the calmodulin inhibitor, calmidazolium, whereas it is acutely activated by NH4+. The homologous erythroid/kidney AE1/SLC4A1 polypeptide is insensitive to these regulators. Each of these AE2 regulatory responses requires the presence of AE2’s C-terminal transmembrane domain (TMD). We have now measured 36Cl− efflux from Xenopus oocytes expressing bi- or tripartite AE2–AE1 chimeras to define TMD subregions in which AE2-specific sequences contribute to acute regulation. The chimeric AE polypeptides were all functional at pHo 7.4, with the sole exception of AE2(1-920)/AE1(613-811)/AE2(1120-1237). Reciprocal exchanges of the large third extracellular loops were without effect. AE2 regulation by pHi, pHo and NH4+ was retained after substitution of C-terminal AE2 amino acids 1120–1237 (including the putative second re-entrant loop, two TM spans and the cytoplasmic tail) with the corresponding AE1 sequence. In contrast, the presence of this AE2 C-terminal sequence was both necessary and sufficient for inhibition by calmidazolium. All other tested TMD substitutions abolished AE2 pHi sensitivity, abolished or severely attenuated sensitivity to pHo and removed sensitivity to NH4+. Loss of AE2 pHi sensitivity was not rescued by co-expression of a complementary AE2 sequence within separate full-length chimeras or AE2 subdomains. Thus, normal regulation of AE2 by pH and other ligands requires AE2-specific sequence from most regions of the AE2 TMD, with the exceptions of the third extracellular loop and a short C-terminal sequence. We conclude that the individual TMD amino acid residues previously identified as influencing acute regulation of AE2 exert that influence within a regulatory structure requiring essential contributions from multiple regions of the AE2 TMD.

A. K. Stewart – One of the best experts on this subject based on the ideXlab platform.

  • Putative re-entrant loop 1 of AE2 transmembrane domain has a major role in acute regulation of anion exchange by pH.
    The Journal of biological chemistry, 2008
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Seth L Alper
    Abstract:

    Normal pH sensitivity of the SLC4A2/AE2 anion exchanger requires transmembrane domain (TMD) amino acid (aa) residues not conserved in the homologous but relatively pH-insensitive SLC4A1/AE1 polypeptide. We tested the hypothesis that the nonconserved aa cluster 1075DKPK1078 within the first putative re-entrant loop (RL1) of AE2 TMD contributes to pH sensor function by studying anion exchange function of AE2 mutants in which these and other RL1 aa were systematically substituted with corresponding RL1 aa from AE1. Regulation of Cl-/Cl- and Cl-/HCO(-)3 exchange by intracellular pH (pHi) or extracellular pH (pHo) was measured as 4,4′-di-isothiocyanatostilbene-2,2′ disulfonic acid-sensitive 36Cl- efflux from Xenopus oocytes. AE2 RL1 mutants 1075AAAQ1078 and 1075AAAQN1079 showed reduced pHi sensitivity and pHo sensitivity was acid-shifted by approximately 1 pH unit. Individual mutants D1075A and P1077A exhibited moderately altered pH sensitivity, whereas a range of substitutions at conserved AE2 Ile-1079 substantially altered sensitivity to pHo and/or pHi. Substitution of the complete AE1 RL1 with AE2 RL1 failed to confer AE2-like pH sensitivity onto AE1. Replacement, however, of AE1 RL1 763SGPGAAAQ770 with AE2 1071VAPGDKPK1078 restored pHi sensitivity to the chimera AE2(1-920)/AE1(613-929) without affecting its low sensitivity to pHo. The results show that acute regulation of AE2 by pH requires RL1 of the TMD. We propose that critical segments of RL1 constitute part of an AE2 pH sensor that, together with residues within the N-terminal half of the TMD, constrain the AE2 polypeptide in a conformation required for regulation of anion exchange by pHi.

  • Acute regulation of mouse AE2 anion exchanger requires isoform-specific amino acid residues from most of the transmembrane domain
    The Journal of Physiology, 2007
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Boris E. Shmukler, Seth L Alper
    Abstract:

    The widely expressed anion exchanger polypeptide AE2/SLC4A2 is acutely inhibited by acidic intracellular (pHi), by acidic extracellular pH (pHo), and by the calmodulin inhibitor, calmidazolium, whereas it is acutely activated by NH4+. The homologous erythroid/kidney AE1/SLC4A1 polypeptide is insensitive to these regulators. Each of these AE2 regulatory responses requires the presence of AE2’s C-terminal transmembrane domain (TMD). We have now measured 36Cl− efflux from Xenopus oocytes expressing bi- or tripartite AE2–AE1 chimeras to define TMD subregions in which AE2-specific sequences contribute to acute regulation. The chimeric AE polypeptides were all functional at pHo 7.4, with the sole exception of AE2(1-920)/AE1(613-811)/AE2(1120-1237). Reciprocal exchanges of the large third extracellular loops were without effect. AE2 regulation by pHi, pHo and NH4+ was retained after substitution of C-terminal AE2 amino acids 1120–1237 (including the putative second re-entrant loop, two TM spans and the cytoplasmic tail) with the corresponding AE1 sequence. In contrast, the presence of this AE2 C-terminal sequence was both necessary and sufficient for inhibition by calmidazolium. All other tested TMD substitutions abolished AE2 pHi sensitivity, abolished or severely attenuated sensitivity to pHo and removed sensitivity to NH4+. Loss of AE2 pHi sensitivity was not rescued by co-expression of a complementary AE2 sequence within separate full-length chimeras or AE2 subdomains. Thus, normal regulation of AE2 by pH and other ligands requires AE2-specific sequence from most regions of the AE2 TMD, with the exceptions of the third extracellular loop and a short C-terminal sequence. We conclude that the individual TMD amino acid residues previously identified as influencing acute regulation of AE2 exert that influence within a regulatory structure requiring essential contributions from multiple regions of the AE2 TMD.

  • Role of nonconserved charged residues of the AE2 transmembrane domain in regulation of anion exchange by pH
    Pflügers Archiv – European Journal of Physiology, 2007
    Co-Authors: A. K. Stewart, C. E. Kurschat, S. L. Alper
    Abstract:

    The ubiquitous AE2/SLC4A2 anion exchexchanger is acutely and independently regulated by intracellular (pH_i) and extracellular pH (pH_o), whereas the closely related AE1/SLC4A1 of the red cell and renal intercalated cell is relatively pH-insensitive. We have investigated the contribution of nonconserved charged residues within the C-terminal transmembrane domain (TMD) of AE2 to regulation by pH through mutation to the corresponding AE1 residues. AE2-mediated Cl^−/Cl^− exchange was measured as 4,4′-di-isothiocyanatostilbene-2,2′-disulfonic acid-sensitive ^36Cl^− efflux from Xenopus oocytes by varying pH_i at constant pH_o, and by varying pH_o at near-constant pH_i. All mutations of nonconserved charged residues of the AE2 TMD yielded functional protein, but mutations of some conserved charged residues (R789E, R1056A, R1134C) reduced or abolished function. Individual mutation of AE2 TMD residues R921, F922, P1077, and R1107 exhibited reduced pH_i sensitivity compared to wt AE2, whereas TMD mutants K1153R, R1155K, R1202L displayed enhanced sensitivity to acidic pH_i. In addition, pH_o sensitivity was significantly acid- shifted when nonconserved AE2 TMD residues E981, K982, and D1075 were individually converted to the corresponding AE1 residues. These results demonstrate that multiple conserved charged residues are important for basal transport function of AE2 and that certain nonconserved charged residues of the AE2 TMD are essential for wild-type regulation of anion exchange by pH_i and pH_o.

C. E. Kurschat – One of the best experts on this subject based on the ideXlab platform.

  • Putative re-entrant loop 1 of AE2 transmembrane domain has a major role in acute regulation of anion exchange by pH.
    The Journal of biological chemistry, 2008
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Seth L Alper
    Abstract:

    Normal pH sensitivity of the SLC4A2/AE2 anion exchanger requires transmembrane domain (TMD) amino acid (aa) residues not conserved in the homologous but relatively pH-insensitive SLC4A1/AE1 polypeptide. We tested the hypothesis that the nonconserved aa cluster 1075DKPK1078 within the first putative re-entrant loop (RL1) of AE2 TMD contributes to pH sensor function by studying anion exchange function of AE2 mutants in which these and other RL1 aa were systematically substituted with corresponding RL1 aa from AE1. Regulation of Cl-/Cl- and Cl-/HCO(-)3 exchange by intracellular pH (pHi) or extracellular pH (pHo) was measured as 4,4′-di-isothiocyanatostilbene-2,2′ disulfonic acid-sensitive 36Cl- efflux from Xenopus oocytes. AE2 RL1 mutants 1075AAAQ1078 and 1075AAAQN1079 showed reduced pHi sensitivity and pHo sensitivity was acid-shifted by approximately 1 pH unit. Individual mutants D1075A and P1077A exhibited moderately altered pH sensitivity, whereas a range of substitutions at conserved AE2 Ile-1079 substantially altered sensitivity to pHo and/or pHi. Substitution of the complete AE1 RL1 with AE2 RL1 failed to confer AE2-like pH sensitivity onto AE1. Replacement, however, of AE1 RL1 763SGPGAAAQ770 with AE2 1071VAPGDKPK1078 restored pHi sensitivity to the chimera AE2(1-920)/AE1(613-929) without affecting its low sensitivity to pHo. The results show that acute regulation of AE2 by pH requires RL1 of the TMD. We propose that critical segments of RL1 constitute part of an AE2 pH sensor that, together with residues within the N-terminal half of the TMD, constrain the AE2 polypeptide in a conformation required for regulation of anion exchange by pHi.

  • Acute regulation of mouse AE2 anion exchanger requires isoform-specific amino acid residues from most of the transmembrane domain
    The Journal of Physiology, 2007
    Co-Authors: A. K. Stewart, C. E. Kurschat, Richard D. Vaughan-jones, Boris E. Shmukler, Seth L Alper
    Abstract:

    The widely expressed anion exchanger polypeptide AE2/SLC4A2 is acutely inhibited by acidic intracellular (pHi), by acidic extracellular pH (pHo), and by the calmodulin inhibitor, calmidazolium, whereas it is acutely activated by NH4+. The homologous erythroid/kidney AE1/SLC4A1 polypeptide is insensitive to these regulators. Each of these AE2 regulatory responses requires the presence of AE2’s C-terminal transmembrane domain (TMD). We have now measured 36Cl− efflux from Xenopus oocytes expressing bi- or tripartite AE2–AE1 chimeras to define TMD subregions in which AE2-specific sequences contribute to acute regulation. The chimeric AE polypeptides were all functional at pHo 7.4, with the sole exception of AE2(1-920)/AE1(613-811)/AE2(1120-1237). Reciprocal exchanges of the large third extracellular loops were without effect. AE2 regulation by pHi, pHo and NH4+ was retained after substitution of C-terminal AE2 amino acids 1120–1237 (including the putative second re-entrant loop, two TM spans and the cytoplasmic tail) with the corresponding AE1 sequence. In contrast, the presence of this AE2 C-terminal sequence was both necessary and sufficient for inhibition by calmidazolium. All other tested TMD substitutions abolished AE2 pHi sensitivity, abolished or severely attenuated sensitivity to pHo and removed sensitivity to NH4+. Loss of AE2 pHi sensitivity was not rescued by co-expression of a complementary AE2 sequence within separate full-length chimeras or AE2 subdomains. Thus, normal regulation of AE2 by pH and other ligands requires AE2-specific sequence from most regions of the AE2 TMD, with the exceptions of the third extracellular loop and a short C-terminal sequence. We conclude that the individual TMD amino acid residues previously identified as influencing acute regulation of AE2 exert that influence within a regulatory structure requiring essential contributions from multiple regions of the AE2 TMD.

  • Role of nonconserved charged residues of the AE2 transmembrane domain in regulation of anion exchange by pH
    Pflügers Archiv – European Journal of Physiology, 2007
    Co-Authors: A. K. Stewart, C. E. Kurschat, S. L. Alper
    Abstract:

    The ubiquitous AE2/SLC4A2 anion exchanger is acutely and independently regulated by intracellular (pH_i) and extracellular pH (pH_o), whereas the closely related AE1/SLC4A1 of the red cell and renal intercalated cell is relatively pH-insensitive. We have investigated the contribution of nonconserved charged residues within the C-terminal transmembrane domain (TMD) of AE2 to regulation by pH through mutation to the corresponding AE1 residues. AE2-mediated Cl^−/Cl^− exchange was measured as 4,4′-di-isothiocyanatostilbene-2,2′-disulfonic acid-sensitive ^36Cl^− efflux from Xenopus oocytes by varying pH_i at constant pH_o, and by varying pH_o at near-constant pH_i. All mutations of nonconserved charged residues of the AE2 TMD yielded functional protein, but mutations of some conserved charged residues (R789E, R1056A, R1134C) reduced or abolished function. Individual mutation of AE2 TMD residues R921, F922, P1077, and R1107 exhibited reduced pH_i sensitivity compared to wt AE2, whereas TMD mutants K1153R, R1155K, R1202L displayed enhanced sensitivity to acidic pH_i. In addition, pH_o sensitivity was significantly acid- shifted when nonconserved AE2 TMD residues E981, K982, and D1075 were individually converted to the corresponding AE1 residues. These results demonstrate that multiple conserved charged residues are important for basal transport function of AE2 and that certain nonconserved charged residues of the AE2 TMD are essential for wild-type regulation of anion exchange by pH_i and pH_o.

Ron R Kopito – One of the best experts on this subject based on the ideXlab platform.

  • functional activation of plasma membrane anion exchangers occurs in a pre golgi compartment
    Journal of Cell Biology, 1993
    Co-Authors: Stephan Ruetz, Ann E Lindsey, Cristina L Ward, Ron R Kopito
    Abstract:

    Folding and oligomerization of most plasma membmembrane glycglycoproteins, including those involved in ion transport, occur in the ER and are frequently required for their exit from this organelle. It is currently unknown, however, where or when in the biosynthetic pathway these proteins become functionally active. AE1 and AE2 are tissue-specific, plasma membrane anion transport proteins. Transient expression of AE2 in a eukaryotic cell line leads to an increase in stilbene inhibitable whole cell 35SO4(2-)-efflux consistent with its function as a plasma membrane anion exchexchanger. No such increased transport activity was observed in AE1 transfectants, despite the fact that the two proteins were synthesized in roughly equal portions. In contrast, both AE1 and AE2 expression resulted in significant increase in Cl-/SO4(2-)-exchange in crude microsomes demonstrating that both AE1 and AE2 cDNAs encode functional proteins. Immunofluorescence staining and pulse-chase labeling experiments revealed that while 60% of AE2 is processed to the cell surface of transfectants, AE1 is restricted to an intracellular compartment and never acquires mature oligosaccharides. Crude microsomes from transfected cells were fractionated into plasma membrane and ER-derived vesicles by con A affinity chromatography. All of the AE1 and approximately half of the cellular AE2 was eluted with the ER vesicles, confirming their intracellular localization. Anion transport measurements on these fractions confirmed that the ER-restricted anion exchangers were functional. We conclude that AE1 and AE2 acquire the ability to mediate anion exchange at an early stage of their biosynthesis, before their exit from the ER.

  • functional differences among nonerythroid anion exchangers expressed in a transfected human cell line
    Journal of Biological Chemistry, 1991
    Co-Authors: Beth S Lee, R B Gunn, Ron R Kopito
    Abstract:

    Abstract A new transient expression system has been developed to investigate the function of anion exchangers in vivo. Human 293 cells were cotransfected with AE2 or AE3 cDNA together with a plasmid encoding a cell surface marker protein. Staining of the cells with antibody directed against a cell surface epitope present in the marker protein permitted the detection of cells expressing functional anion exchangers. Intracellular pH (pHi) recording in individual transfectants loaded with the fluorescent pHi indicator, 2′,7′-bis(carboxyethyl)-5,6-carboxyfluorescein, was used to determine the flux of HCO3- as a measure of Cl-/HCO3- exchange activity. Cells expressing either anion exchexchanger displayed significantly enhanced Cl-/HCO3- exchange activity compared with controls expressing only the marker. Transfection with either anion exchexchanger or with control plasmid resulted in altered intrinsic buffering capacity profiles compared with untransfected controls. Expression of either AE2 or AE3 did not result in changes in resting pHi. The activities of both AE2 and AE3 were stimulated at alkaline pHi, suggesting that an internal protonation site in AE2 and AE3 may regulate their activities. Both exchangers were inhibited reversibly and irreversibly by the anion 4,4′-diisothiocyanostilbene-2,2′-disulfonate with IC50 values of 142 and 0.43 microM for AE2 and AE3, respectively. These data indicate that structural differences in these highly conserved anion exchangers give rise to differences in affinities at the external anion binding site.

Iva Juranovic Cindric – One of the best experts on this subject based on the ideXlab platform.

  • determination of trace elements in olive oil by icp Aes and eta aas a pilot study on the geographical characterization
    Microchemical Journal, 2005
    Co-Authors: Michaela Zeiner, Ilse Steffan, Iva Juranovic Cindric
    Abstract:

    Abstract The determination of trace elements in edible oils is important because of both the metabolic role of metals and possibilities for adulteration detection and oil characterization. The most commonly used techniques for the determination of metals in oil samples are inductively coupled plasma atomic emission specspectrometry (ICP-Aes) and atomic absorption specspectrometry (AAS). For this study, a microwave assisted decomposition of the olive oil in closed vessels using a mixture of nitric acid and hydrogen peroxide was applied as sample preparation. The low achievable LODs enable the determination by ICP-Aes of even very low concentrations of most elements of interest. The proposed ICP-Aes method permits the determination of Ca, Fe, Mg, Na, and Zn in olive oils. Elements present in small amounts (Al, Co, Cu, K, Mn, Ni) were measured by ETA-AAS in the same sample digest. The concentrations of Al, Co, Cu, K, Mn, and Ni were in the range from 0.15 to 1.5 μg/g and differ according to the geographical origin of the oils. For the amounts of Fe, Mg, Na, and Zn in the samples, no significant differences according to the geographical origin of the oils could be observed, the mean concentrations being 15.31, 3.26, 33.10, and 3.39 μg/g, respectively. The Ca content varies in the range of 1.3 to 9.0 μg/g. The dependency of the trace elemental content of olive oils on their geographical origin can be used for their local characterization.