| 1. |
- Mim, Carsten, et al.
(författare)
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Structure versus function : Are new conformations of pannexin 1 yet to be resolved?
- 2021
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Ingår i: The Journal of General Physiology. - : Rockefeller University Press. - 0022-1295 .- 1540-7748. ; 153:5
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Tidskriftsartikel (refereegranskat)abstract
- Pannexin 1 (Panx1) plays a decisive role in multiple physiological and pathological settings, including oxygen delivery to tissues, mucociliary clearance in airways, sepsis, neuropathic pain, and epilepsy. It is widely accepted that Panx1 exerts its role in the context of purinergic signaling by providing a transmembrane pathway for ATP. However, under certain conditions, Panx1 can also act as a highly selective membrane channel for chloride ions without ATP permeability. A recent flurry of publications has provided structural information about the Panx1 channel. However, while these structures are consistent with a chloride selective channel, none show a conformation with strong support for the ATP release function of Panx1. In this Viewpoint, we critically assess the existing evidence for the function and structure of the Panx1 channel and conclude that the structure corresponding to the ATP permeation pathway is yet to be determined. We also list a set of additional topics needing attention and propose ways to attain the large-pore, ATP-permeable conformation of the Panx1 channel.
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| 2. |
- Wang, Zuoneng, et al.
(författare)
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Coming of Age: Cryo-Electron Tomography as a Versatile Tool to Generate High-Resolution Structures at Cellular/Biological Interfaces
- 2021
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Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 22:12, s. 6177-6177
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Forskningsöversikt (övrigt vetenskapligt/konstnärligt)abstract
- Over the last few years, cryo electron microscopy has become the most important methodin structural biology. While 80% of deposited maps are from single particle analysis, electrontomography has grown to become the second most important method. In particular sub-tomogramaveraging has matured as a method, delivering structures between 2 and 5 Å from complexes in cellsas well as in vitro complexes. While this resolution range is not standard, novel developments pointtoward a promising future. Here, we provide a guide for the workflow from sample to structure togain insight into this emerging field.
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| 3. |
- Wang, Zuoneng, et al.
(författare)
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CryoEM reveals BIN1 (isoform 8) does not bind to single actinfilaments in vitro
- 2021
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Ingår i: microPublication biology. - : Caltech Library. - 2578-9430.
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Cells change their appearance by a concerted action of the cytoskeleton and the plasma membrane. The machinery thatbends the membrane includes Bin/Amphiphysin/Rvs (BAR) domain proteins. Recently BAR domain proteins garneredattention as actin regulators, either by recruiting actin regulating proteins or through binding to actin directly. BIN1 (animportant protein in Alzheimer’s Disease, heart disease and cancer) is one of the few BAR proteins that bind to actindirectly. Here, we imaged a complex of BIN1 and actin with cryoEM. Our results reveal that BIN1 cannot be found onsingle actin filaments.
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| 4. |
- Wang, Zuoneng, 1991-
(författare)
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Structure studies of membrane associated proteins by transmission electron microscopy
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cell membranes need to change their shapes during many cellular processeslike protein trafficking, cytokinesis and membrane homeostasis. The lattershuttles lipids, synthesized in the endoplasmic reticulum, to all membranouscompartments. Bin/Amphiphysin/Rvs (BAR) proteins are peripheralmembrane proteins (PMP) and play an important role in sculpturingmembranes and in the regulation of actin dynamics. Cryo-electronmicroscopy (cryoEM) has emerged as a powerful tool to visualize proteinsat the membrane interface. Here, we employed transmission electronmicroscopy and other biophysical methods to elucidate how BAR domainproteins steer processes at the membrane.In this work we studied the BAR protein bridging integrator 1 (BIN1), whichhas an established role in cancer, Alzheimer’s disease and skeletalmyopathies. To obtain information about BIN1’s interaction with themembrane in near native environments, we used artificial lipid systems suchas liposomes and lipids nanotubes.First, we have shown that electrostatic interactions are more important forBIN1 when binding to membranes with low curvature. At high curvature,binding is likely driven by non-polar interactions. The formation ofinvaginations (or tubules) is regulated by the composition of negativecharged lipids in membrane bilayer or electrostatic residues on the BARdomain. Therefore electrostatic interactions regulate recruitment andcrowding of BIN1; and consequently membrane deformation.Second, we clarified BIN1’s role in actin dynamics. CryoEM reveals that themuscular BIN1 isoform does not bind to single actin filaments, althoughBIN1 can be co-sedimented with actin after polymerization of actin. Thisimplies that BIN1 rather bundles actin than decorates single filaments.Third, we explored a strategy to purify an aggregation prone BAR protein.Aggregation is a property common in Peripheral Membrane Proteins. Thenovel NT* tag is derived from a spider silk protein and was reported to be apromising fusion tag for protein purification. We showed that the NT* tagimproves the solubility and reduces the aggregation of the BAR proteinFAM92A1. The activity of purified FAM92A1-NT* was verified bynegative stain EM.IIFourth, we were interested in the regulation of the lipid metabolism. PyruvateCarboxylase (PC) is a pivotal enzyme to generate lipid precursors. Cellbiological assays identified a long non-coding (lnc) RNA that regulates theactivity of PC. We studied the interaction between the lnc RNA and PC bybiophysical techniques. Size exclusion chromatography confirmed thepresence lncRNA-PC complex in vitro.
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