Threonine 67 is a key component in the coupling of the NSS amino acid transporter KAAT1

被引:4
|
作者
Giovanola, M. [1 ]
Vollero, A. [2 ,4 ]
Cinquetti, R. [2 ]
Bossi, E. [2 ]
Forrest, L. R. [3 ]
Di Cairano, E. S. [1 ]
Castagna, M. [1 ]
机构
[1] Univ Milan, Dept Pharmacol & Biomol Sci, Via Trentacoste 2, I-20134 Milan, Italy
[2] Univ Insubria, Dept Biotechnol & Life Sci, Via JH Dunant 3, I-21100 Varese, Italy
[3] NINDS, Computat Struct Biol Sect, NIH, 35 Convent Dr, Bethesda, MD 20892 USA
[4] IRCCS Fdn Salvatore Maugeri, Mol Cardiol, Via S Maugeri 10, I-27100 Pavia, Italy
来源
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES | 2018年 / 1860卷 / 05期
关键词
NSS family; Structure-function analysis; coupling mechanism; Amino acid uptake; Ion selectivity; Xenopus laevis oocytes; Homology model; SIDE-CHAIN CONFORMATIONS; XENOPUS-LAEVIS OOCYTES; X-RAY STRUCTURES; DOPAMINE TRANSPORTER; COTRANSPORTER KAAT1; NEUROTRANSMITTER TRANSPORTERS; SUBSTRATE SELECTIVITY; SEROTONIN TRANSPORTER; MOLECULAR PHYSIOLOGY; SODIUM SYMPORTERS;
D O I
10.1016/j.bbamem.2018.01.020
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The crystallizations of the prokaryotic LeuT and of the eukaryotic DAT and SERT transporters represent important steps forward in the comprehension of the molecular physiology of Neurotransmitter: Sodium Symporters, although the molecular determinants of the coupling mechanism and of ion selectivity still remain to be fully elucidated. The insect NSS homologue KAAT1 exhibits unusual physiological features, such as the ability to use K+ as the driver ion, weak chloride dependence, and the ability of the driver ion to influence the substrate selectivity; these characteristics can help to define the molecular determinants of NSS function. Two non-conserved residues are present in the putative sodium binding sites of KAAT1: Ala 66, corresponding to Gly 20 in the Na2 site of LeuT, and Ser 68, corresponding to Ala 22 in the Nal site. Thr 67 appears also to be significant since it is not conserved among NSS members, is present as threonine only in KAAT1 and in the paralogue CAATCH1 and, according to LeuT structure, is close to the amino acid binding site. Mutants of these residues were functionally characterized in Xenopus oocytes. The T67Y mutant exhibited uptake activity comparable to that of the wild type, but fully chloride-independent and with enhanced stereoselectivity. Interestingly, although dependent on the presence of sodium, the mutant showed reduced transport-associated currents, indicating uncoupling of the driver ion and amino acid fluxes. Thr 67 therefore appears to be a key component in the coupling mechanism, participating in a network that influences the cotransport of Na+ and the amino acid.
引用
收藏
页码:1179 / 1186
页数:8
相关论文
共 50 条
  • [1] Role of a conserved glycine triplet in the NSS amino acid transporter KAAT1
    Giovanola, M.
    D'Antoni, F.
    Santacroce, M.
    Mari, S. A.
    Cherubino, F.
    Bossi, E.
    Sacchi, V. F.
    Castagna, M.
    BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES, 2012, 1818 (07): : 1737 - 1744
  • [2] Ionic selectivity of the coupled and uncoupled currents carried by the amino acid transporter KAAT1
    Bossi, E
    Sacchi, VF
    Peres, A
    PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 1999, 438 (06): : 788 - 796
  • [3] Ionic selectivity of the coupled and uncoupled currents carried by the amino acid transporter KAAT1
    E. Bossi
    V. F. Sacchi
    A. Peres
    Pflügers Archiv, 1999, 438 (6): : 788 - 796
  • [4] Inhibition of the lepidopteran amino acid co-transporter KAAT1 by phenylglyoxal: role of arginine 76
    Castagna, M
    Vincenti, S
    Marciani, P
    Sacchi, VF
    INSECT MOLECULAR BIOLOGY, 2002, 11 (04) : 283 - 289
  • [5] Structural and functional basis of amino acid specificity in the invertebrate cotransporter KAAT1
    Miszner, Andreea
    Peres, Antonio
    Castagna, Michela
    Bette, Sara
    Giovannardi, Stefano
    Cherubino, Francesca
    Bossi, Elena
    JOURNAL OF PHYSIOLOGY-LONDON, 2007, 581 (03): : 899 - 913
  • [6] Ion binding and permeation through the lepidopteran amino acid transporter KAAT1 expressed in Xenopus oocytes
    Bossi, E
    Centinaio, E
    Castagna, M
    Giovannardi, S
    Vincenti, S
    Sacchi, VF
    Peres, A
    JOURNAL OF PHYSIOLOGY-LONDON, 1999, 515 (03): : 729 - 742
  • [7] Aspartate 338 contributes to the cationic specificity and to driver-amino acid coupling in the insect cotransporter KAAT1
    S. A. Mari
    A. Soragna
    M. Castagna
    E. Bossi
    A. Peres
    V. F. Sacchi
    Cellular and Molecular Life Sciences CMLS, 2004, 61 : 243 - 256
  • [8] Immunocytochemical localization of the amino acid co-transporter KAAT1 and neuromodulators in the midgut of larval Manduca sexta.
    Sanderlin, A. G.
    Rose, E. K.
    Yeoh, A. J.
    Gillen, C. M.
    Itagaki, H.
    INTEGRATIVE AND COMPARATIVE BIOLOGY, 2013, 53 : E365 - E365
  • [9] Oligomeric structure of the neutral amino acid transporters KAAT1 and CAATCH1
    Bossi, Elena
    Soragna, Andrea
    Miszner, Andreea
    Giovannardi, Stefano
    Frangione, Valeria
    Peres, Antonio
    AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY, 2007, 292 (04): : C1379 - C1387
  • [10] Expression cloning of a K+-coupled amino acid transporter (KAAT1) from lepidopteran larval intestine.
    Castagna, M
    Shayakul, C
    Trotti, D
    Sacchi, VF
    Harvey, WR
    Hediger, MA
    FASEB JOURNAL, 1997, 11 (03): : 181 - 181