Ion channels and their molecular environments – Glimpses and insights from functional proteomics

Schulte, Uwe; Müller, Christoph Michael; Fakler, Bernd

doi:10.1016/j.semcdb.2010.09.015

Cited by 29 publications

(35 citation statements)

References 125 publications

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“…The diversity in electrical properties among different cell types or states is largely defined by expression and subunit composition of individual ion channel pore-forming principle subunits and often tissue-or cell type-specific regulatory subunits (1,2). The large conductance, calcium-and voltage-activated potassium channel (BK, also termed as BK Ca , Maxi-K, K Ca 1.1, or Slo1) is a unique member of the potassium channel family, which is dually activated by membrane depolarization and elevated intracellular free Ca 2+ ([Ca 2+ ] i ), playing a powerful integrative role in the regulation of cellular excitability and calcium signaling (3).…”

Section: +mentioning

confidence: 99%

BK potassium channel modulation by leucine-rich repeat-containing proteins

Yan

Aldrich²

2012

Proc. Natl. Acad. Sci. U.S.A.

193

288

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Molecular diversity of ion channel structure and function underlies variability in electrical signaling in nerve, muscle, and nonexcitable cells. Regulation by variable auxiliary subunits is a major mechanism to generate tissue-or cell-specific diversity of ion channel function. Mammalian large-conductance, voltage-and calcium-activated potassium channels (BK, K Ca 1.1) are ubiquitously expressed with diverse functions in different tissues or cell types, consisting of the pore-forming, voltage-and Ca 2+-sensing α-subunits (BKα), either alone or together with the tissue-specific auxiliary β-subunits (β1-β4). We recently identified a leucine-rich repeat (LRR)-containing membrane protein, LRRC26, as a BK channel auxiliary subunit, which causes an unprecedented large negative shift (∼140 mV) in voltage dependence of channel activation. Here we report a group of LRRC26 paralogous proteins, LRRC52, LRRC55, and LRRC38 that potentially function as LRRC26-type auxiliary subunits of BK channels. LRRC52, LRRC55, and LRRC38 produce a marked shift in the BK channel's voltage dependence of activation in the hyperpolarizing direction by ∼100 mV, 50 mV, and 20 mV, respectively, in the absence of calcium. They along with LRRC26 show distinct expression in different human tissues: LRRC26 and LRRC38 mainly in secretory glands, LRRC52 in testis, and LRRC55 in brain. LRRC26 and its paralogs are structurally and functionally distinct from the β-subunits and we designate them as a γ family of the BK channel auxiliary proteins, which potentially regulate the channel's gating properties over a spectrum of different tissues or cell types.accessory protein | patch clamp | signal peptide | cotranslational expression

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Section: +mentioning

confidence: 99%

BK potassium channel modulation by leucine-rich repeat-containing proteins

Yan

Aldrich²

2012

Proc. Natl. Acad. Sci. U.S.A.

193

288

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“…Antibody-based affinity purification (AP) 1 of protein assemblies from biological samples followed by mass spectrometric analysis represents an increasingly popular approach for identification of protein-protein interactions (AP-MS) (1)(2)(3). Despite the exquisitely high and specific enrichment theoretically obtainable with antibodies (Abs), this approach faces a number of technical and intrinsic challenges in practice.…”

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confidence: 99%

Extending the Dynamic Range of Label-free Mass Spectrometric Quantification of Affinity Purifications

Bildl

Haupt

Müller

et al. 2012

Molecular & Cellular Proteomics

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Affinity purification (AP) of protein complexes combined with LC-MS/MS analysis is the current method of choice for identification of protein-protein interactions. Their interpretation with respect to significance, specificity, and selectivity requires quantification methods coping with enrichment factors of more than 1000-fold, variable amounts of total protein, and low abundant, unlabeled samples. We used standardized samples (0.1-1000 fmol) measured on high resolution hybrid linear ion trap instruments (LTQ-FT/Orbitrap) to characterize and improve linearity and dynamic range of label-free approaches. Quantification based on spectral counts was limited by saturation and ion suppression effects with samples exceeding 100 ng of protein, depending on the instrument setup. In contrast, signal intensities of peptides (peak volumes) selected by a novel correlation-based method (TopCorr-PV) were linear over at least 4 orders of magnitude and allowed for accurate relative quantification of standard proteins spiked into a complex protein background. Application of this procedure to APs of the voltage-gated potassium channel Kv1.1 as a model membrane protein complex unambiguously identified the whole set of known interaction partners together with novel candidates. In addition to discriminating these proteins from background, we could determine efficiency, cross-reactivities, and selection biases of the used purification antibodies. The enhanced dynamic range of the developed quantification procedure appears well suited for sensitive identification of specific protein-protein interactions, detection of antibody-related artifacts, and optimization of AP conditions. Molecular & Cellular Proteomics 11: 10.1074/mcp.M111.007955, 1-12, 2012.Antibody-based affinity purification (AP) 1 of protein assemblies from biological samples followed by mass spectrometric analysis represents an increasingly popular approach for identification of protein-protein interactions (AP-MS) (1-3). Despite the exquisitely high and specific enrichment theoretically obtainable with antibodies (Abs), this approach faces a number of technical and intrinsic challenges in practice. Target protein complexes typically suffer from poor solubility, instability, and low abundance, particularly when associated with lipid membranes. Moreover, various antibody-related properties such as target selectivity, cross-reactivity, and interference with protein-protein interactions may lead to falsepositive and false-negative results (4). Finally, biological protein-protein interactions may have a more dynamic character, may depend on regulated modifications, or may involve rare protein partners. Together, these effects lead to a significant reduction of AP signal to noise, i.e. low co-enrichment efficiency of interaction partners and significant overlap with background or nonspecific proteins.Classically, AP specificity has been addressed by visualization of purified proteins on one-or two-dimensional gels and comparison of band patterns or spots with those obtained in...

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“…Super resolution microscopy approaches with temporal resolution >30 Hz, such as sptPALM, UPAINT or SPT, 55,80,90 will help to determine the temporal molecular dynamics and organization of ion channel complexes and even enable us to observe those proteins during channel activity. Keeping in mind the astonishing number of potential interaction candidates, as exemplified for AMPA receptors, VGCCs and potassium channels, 36,85,89,100,120,121 the combination of single molecular imaging approaches, in conjunction with functional readouts, will facilitate the investigation of multiple facets of the dynamic contribution of ion channels, as both ion conductors and signaling complexes, for the plasticity, formation and maintenance of neuronal connectivity, and will open new vistas for the identification of pathological mechanisms of neurological disorders on the molecular level. …”

Section: Potassium Channelsmentioning

confidence: 99%

Surface dynamics of voltage-gated ion channels

et al. 2016

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Neurons encode information in fast changes of the membrane potential, and thus electrical membrane properties are critically important for the integration and processing of synaptic inputs by a neuron. These electrical properties are largely determined by ion channels embedded in the membrane. The distribution of most ion channels in the membrane is not spatially uniform: they undergo activity-driven changes in the range of minutes to days. Even in the range of milliseconds, the composition and topology of ion channels are not static but engage in highly dynamic processes including stochastic or activity-dependent transient association of the pore-forming and auxiliary subunits, lateral diffusion, as well as clustering of different channels. In this review we briefly discuss the potential impact of mobile sodium, calcium and potassium ion channels and the functional significance of this for individual neurons and neuronal networks.

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Ion channels and their molecular environments – Glimpses and insights from functional proteomics

Cited by 29 publications

References 125 publications

BK potassium channel modulation by leucine-rich repeat-containing proteins

BK potassium channel modulation by leucine-rich repeat-containing proteins

Extending the Dynamic Range of Label-free Mass Spectrometric Quantification of Affinity Purifications

Surface dynamics of voltage-gated ion channels

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