Proteomic analyses of native brain K<sub>V</sub>4.2 channel complexes

Marionneau, Céline; LeDuc, Richard D.; Rohrs, Henry W.; Link, Andrew J.; Townsend, R. Reid; Nerbonne, Jeanne M.

doi:10.4161/chan.3.4.9553

Cited by 32 publications

(55 citation statements)

References 43 publications

(61 reference statements)

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“…DPP6 in brain and demonstrated that DPP6 immunoprecipitates with Kv4.2 from adult rat and mouse brain (14,15,48), consistent with suggestions that DPP6 is a component of native Kv4.2-encoded I A channel complexes (13,30). Consistent with these findings and with a role for DPP6 in the generation of I A channels in cortical neurons, Western blot analysis revealed robust expression of DPP6 in wild type adult mouse visual cortex (Fig.…”

Section: Cortical Dpp6 Expression Is Unaffected By Loss Of Kv42 or Ksupporting

confidence: 87%

“…Increasing evidence suggests that native neuronal I A channels function in macromolecular protein complexes comprising Kv4 poreforming (␣) subunits, together with Kv4 channel accessory subunits, including the K ϩ channel interacting proteins (KChIP1-4), dipeptidyl peptidases 6 and 10 (DPP6 and DPP10), and additional regulatory and scaffolding proteins (13)(14)(15)(16)(17)(18)(19). Interestingly, recent studies have identified DPP6 and DPP10 as genes associated with autism (20), as well as amyotrophic lateral sclerosis (21,22), although the pathogenic mechanisms remain to be elucidated.…”

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confidence: 99%

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Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

Foeger¹,

Norris²,

Wren

et al. 2012

Journal of Biological Chemistry

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Section: Cortical Dpp6 Expression Is Unaffected By Loss Of Kv42 or Ksupporting

confidence: 87%

mentioning

confidence: 99%

Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

Foeger¹,

Norris²,

Wren

et al. 2012

Journal of Biological Chemistry

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“…S4) and showed several interesting features of the respective Ab: proteins fell into categories (quadrants) of specifically co-enriched, cross-reactive, and different types of background-related proteins; the formation of distinct clusters rather than continuous distributions suggests that proteins have been captured not independently from each other but as defined assemblies. Although distribution patterns and derived specificity thresholds may vary with different Abs, targets, source membranes, and AP conditions, the large number of nonspecific proteins compared with candidate Kv1.1-interactors is rather typical (11,16,19). This emphasizes the need for both IgG and knockout controls to obtain true positive AP-MS results.…”

Section: Discussionmentioning

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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“…35 The tandem spectra were used to search protein databases with Mascot software (Matrix Science, London, UK; version 2.1.1.0). The nr_20080708 database (selected for Homo sapiens, 127,310 entries) was searched using trypsin as the endoprotease, a parent mass tolerance of 20 ppm, and a fragment ion mass tolerance of 0.7 Da.…”

Section: Database Searchingmentioning

confidence: 99%

Inhibition of Lens Photodamage by UV-Absorbing Contact Lenses

Andley

Malone²,

Townsend

2011

Invest. Ophthalmol. Vis. Sci.

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Proteomic analyses of native brain K_V4.2 channel complexes

Cited by 32 publications

References 43 publications

Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

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

Inhibition of Lens Photodamage by UV-Absorbing Contact Lenses

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Proteomic analyses of native brain KV4.2 channel complexes

Cited by 32 publications

References 43 publications

Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

Augmentation of Kv4.2-encoded Currents by Accessory Dipeptidyl Peptidase 6 and 10 Subunits Reflects Selective Cell Surface Kv4.2 Protein Stabilization

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

Inhibition of Lens Photodamage by UV-Absorbing Contact Lenses

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Proteomic analyses of native brain K_V4.2 channel complexes