Phospho-regulation, nucleotide binding and ion access control in potassium-chloride cotransporters.
Phospho-regulation, nucleotide binding and ion access control in potassium-chloride cotransporters.
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DOI：
10.15252/embj.2020107294
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发表时间：
2021-07-15
期刊：
影响因子：
--
通讯作者：
Dürr KL
中科院分区：
文献类型：
--
作者：
Chi G;Ebenhoch R;Man H;Tang H;Tremblay LE;Reggiano G;Qiu X;Bohstedt T;Liko I;Almeida FG;Garneau AP;Wang D;McKinley G;Moreau CP;Bountra KD;Abrusci P;Mukhopadhyay SMM;Fernandez-Cid A;Slimani S;Lavoie JL;Burgess-Brown NA;Tehan B;DiMaio F;Jazayeri A;Isenring P;Robinson CV;Dürr KL
钾偶联氯离子转运体（KCCs）在调节细胞体积和细胞内氯离子浓度中起着至关重要的作用。它们在等渗条件下通过位于细胞质末端的磷酸化调控位点被特异性地抑制。低渗细胞肿胀引起的抑制性磷酸化降低刺激转运活性，这一调节过程的功能障碍与多种人类疾病有关。在这里，我们展示了人类KCC3b和KCC1的低温电镜结构，揭示了N端和C端磷酸化调控的结构决定因素。我们发现，模拟磷酸的KCC3b处于向内的状态，在这种状态下，细胞内的离子通道被与N端广泛接触所阻断。在另一个等压转运活性增加的突变体KCC1Δ19中，这种结构域间的相互作用是不存在的，可能是由于KCC1 N端有一个独特的磷酸化调控位点。此外，我们在大C端结构域中绘制了额外的磷酸化位点以及以前未知的ATP/ADP结合口袋，并显示了腺嘌呤核苷酸增强了其他CCCs的热稳定性。这些发现为kcc的复杂调控提供了根本性的新见解，并可能为药物开发提供创新策略。低温电镜结构揭示了磷酸化依赖的蛋白质动力学和ATP/ADP结合的人类KCC膜转运蛋白。
Potassium‐coupled chloride transporters (KCCs) play crucial roles in regulating cell volume and intracellular chloride concentration. They are characteristically inhibited under isotonic conditions via phospho‐regulatory sites located within the cytoplasmic termini. Decreased inhibitory phosphorylation in response to hypotonic cell swelling stimulates transport activity, and dysfunction of this regulatory process has been associated with various human diseases. Here, we present cryo‐EM structures of human KCC3b and KCC1, revealing structural determinants for phospho‐regulation in both N‐ and C‐termini. We show that phospho‐mimetic KCC3b is arrested in an inward‐facing state in which intracellular ion access is blocked by extensive contacts with the N‐terminus. In another mutant with increased isotonic transport activity, KCC1Δ19, this interdomain interaction is absent, likely due to a unique phospho‐regulatory site in the KCC1 N‐terminus. Furthermore, we map additional phosphorylation sites as well as a previously unknown ATP/ADP‐binding pocket in the large C‐terminal domain and show enhanced thermal stabilization of other CCCs by adenine nucleotides. These findings provide fundamentally new insights into the complex regulation of KCCs and may unlock innovative strategies for drug development. Cryo‐EM structures uncover phosphorylation‐dependent protein dynamics and ATP/ADP binding of human KCC membrane transporters.
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影响因子：
8.6
作者：
Jelcic M;Wang K;Hui KL;Cai XC;Enyedi B;Luo M;Niethammer P
通讯作者：
Niethammer P
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
2
作者：
Jin, Sheng Chih;Furey, Charuta G.;Kahle, Kristopher T.
通讯作者：
Kahle, Kristopher T.
KCC3在维持周围神经纤维的细胞体积中的作用。
影响因子：
4.2
作者：
Flores B;Schornak CC;Delpire E
通讯作者：
Delpire E
影响因子：
5.3
作者：
Adragna NC;Ravilla NB;Lauf PK;Begum G;Khanna AR;Sun D;Kahle KT
通讯作者：
Kahle KT