| 1. |
- Törnroth-Horsefield, Susanna, 1973, et al.
(author)
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Aquaporin gating
- 2007
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In: Journal of Biomolecular Structure & Dynamics. ; 24:6, s. 719-721
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Journal article (peer-reviewed)
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| 2. |
- Törnroth-Horsefield, Susanna, 1973, et al.
(author)
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Crystal Structure of AcrB in Complex with a Single Transmembrane Subunit Reveals Another Twist.
- 2007
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In: Structure (London, England : 1993). - : Elsevier BV. - 0969-2126 .- 1878-4186. ; 15:12, s. 1663-73
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Journal article (peer-reviewed)abstract
- Bacterial drug resistance is a serious concern for human health. Multidrug efflux pumps export a broad variety of substrates out of the cell and thereby convey resistance to the host. In Escherichia coli, the AcrB:AcrA:TolC efflux complex forms a principal transporter for which structures of the individual component proteins have been determined in isolation. Here, we present the X-ray structure of AcrB in complex with a single transmembrane protein, assigned by mass spectrometry as YajC. A specific rotation of the periplasmic porter domain of AcrB is also revealed, consistent with the hypothesized "twist-to-open" mechanism for TolC activation. Growth experiments with yajc-deleted E. coli reveal a modest increase in the organism's susceptibility to beta-lactam antibiotics, but this effect could not conclusively be attributed to the loss of interactions between YajC and AcrB.
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| 3. |
- Adawy, Alaa, et al.
(author)
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High Resolution Protein Crystals Using an Efficient Convection-Free Geometry
- 2013
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In: Crystal Growth & Design. - : American Chemical Society (ACS). - 1528-7483 .- 1528-7505. ; 13:2, s. 775-781
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Journal article (peer-reviewed)abstract
- Macromolecular crystallography is the most direct and accurate approach to determine the three-dimensional structure of biological macromolecules. The growth of high quality single crystals, yielding diffraction to the highest X-ray resolution, remains a bottleneck in this methodology. Here we show that through a modification of the batch crystallization method, an entirely convection-free crystallization environment is achieved, which enhances the purity and crystallinity of protein crystals. This is accomplished by using an upside-down geometry, where crystals grow at the “ceiling” of a growth-cell completely filled with the crystallization solution. The “ceiling crystals” experience the same diffusion-limited conditions as in space microgravity experiments. The new method was tested on bovine insulin and two hen egg-white lysozyme polymorphs. In all cases, ceiling crystals diffracted X-rays to resolution limits beyond that for other methods using similar crystallization conditions without further optimization. In addition, we demonstrate that the ceiling crystallization method leads to crystals with much lower impurity incorporation.
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| 4. |
- Al-Jubair, Tamim, et al.
(author)
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Characterization of human aquaporin protein-protein interactions using microscale thermophoresis (MST)
- 2022
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In: STAR Protocols. - : Elsevier BV. - 2666-1667. ; 3:2
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Journal article (peer-reviewed)abstract
- Aquaporin water channels (AQPs) are membrane proteins that maintain cellular water homeostasis. The interactions between human AQPs and other proteins play crucial roles in AQP regulation by both gating and trafficking. Here, we describe a protocol for characterizing the interaction between a human AQP and a soluble interaction partner using microscale thermophoresis (MST). MST has the advantage of low sample consumption and high detergent compatibility enabling AQP protein-protein interaction investigation with a high level of control of components and environment. For complete details on the use and execution of this protocol, please refer to Kitchen et al. (2020) and Roche et al. (2017).
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| 5. |
- Al-Jubair, Tamim, et al.
(author)
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High-yield overproduction and purification of human aquaporins from Pichia pastoris
- 2022
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In: STAR Protocols. - : Elsevier BV. - 2666-1667. ; 3:2
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Journal article (peer-reviewed)abstract
- Aquaporins (AQPs) are membrane-bound water channels that play crucial roles in maintaining the water homeostasis of the human body. Here, we present a protocol for high-yield recombinant expression of human AQPs in the methylotropic yeast Pichia pastoris and subsequent AQP purification. The protocol typically yields 1–5 mg AQP per g of yeast cell at >95% purity and is compatible with any membrane protein cloned into Pichia pastoris, although expression levels may vary. For complete details on the use and execution of this protocol, please refer to Kitchen et al. (2020) and Frick et al. (2014).
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| 6. |
- Chivasso, Clara, et al.
(author)
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Ezrin is a novel protein partner of aquaporin-5 in human salivary glands and shows altered expression and cellular localization in sjögren’s syndrome
- 2021
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In: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 22:17
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Journal article (peer-reviewed)abstract
- Sjögren’s syndrome (SS) is an exocrinopathy characterized by the hypofunction of salivary glands (SGs). Aquaporin-5 (AQP5); a water channel involved in saliva formation; is aberrantly dis-tributed in SS SG acini and contributes to glandular dysfunction. We aimed to investigate the role of ezrin in AQP5 mislocalization in SS SGs. The AQP5–ezrin interaction was assessed by immuno-precipitation and proteome analysis and by proximity ligation assay in immortalized human SG cells. We demonstrated, for the first time, an interaction between ezrin and AQP5. A model of the complex was derived by computer modeling and in silico docking; suggesting that AQP5 interacts with the ezrin FERM-domain via its C-terminus. The interaction was also investigated in human minor salivary gland (hMSG) acini from SS patients (SICCA-SS); showing that AQP5–ezrin complexes were absent or mislocalized to the basolateral side of SG acini rather than the apical region compared to controls (SICCA-NS). Furthermore, in SICCA-SS hMSG acinar cells, ezrin immunore-activity was decreased at the acinar apical region and higher at basal or lateral regions, accounting for altered AQP5–ezrin co-localization. Our data reveal that AQP5–ezrin interactions in human SGs could be involved in the regulation of AQP5 trafficking and may contribute to AQP5-altered localization in SS patients.
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| 7. |
- Chivasso, Clara, et al.
(author)
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Unraveling human aqp5-pip molecular interaction and effect on aqp5 salivary glands localization in ss patients
- 2021
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In: Cells. - : MDPI AG. - 2073-4409. ; 10:8
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Journal article (peer-reviewed)abstract
- Saliva secretion requires effective translocation of aquaporin 5 (AQP5) water channel to the salivary glands (SGs) acinar apical membrane. Patients with Sjögren’s syndrome (SS) display abnormal AQP5 localization within acinar cells from SGs that correlate with sicca manifestation and glands hypofunction. Several proteins such as Prolactin-inducible protein (PIP) may regulate AQP5 trafficking as observed in lacrimal glands from mice. However, the role of the AQP5-PIP complex remains poorly understood. In the present study, we show that PIP interacts with AQP5 in vitro and in mice as well as in human SGs and that PIP misexpression correlates with an altered AQP5 distribution at the acinar apical membrane in PIP knockout mice and SS hMSG. Furthermore, our data show that the protein-protein interaction involves the AQP5 C-terminus and the N-terminal of PIP (one molecule of PIP per AQP5 tetramer). In conclusion, our findings highlight for the first time the role of PIP as a protein controlling AQP5 localization in human salivary glands but extend beyond due to the PIP-AQP5 interaction described in lung and breast cancers.
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| 8. |
- Frick, Anna, 1982, et al.
(author)
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Mercury increases water permeability of a plant aquaporin through a non-cysteine-related mechanism
- 2013
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In: Biochemical Journal. - : Portland Press Ltd.. - 0264-6021 .- 1470-8728. ; 454:pt 3, s. 491-499
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Journal article (peer-reviewed)abstract
- Water transport across cellular membranes is mediated by a family of membrane proteins known as AQPs (aquaporins). AQPs were first discovered on the basis of their ability to be inhibited by mercurial compounds, an experiment which has followed the AQP field ever since. Although mercury inhibition is most common, many AQPs are mercury insensitive. In plants, regulation of AQPs is important in order to cope with environmental changes. Plant plasma membrane AQPs are known to be gated by phosphorylation, pH and Ca2+. We have previously solved the structure of the spinach AQP SoPIP2;1 (Spinacia oleracea plasma membrane intrinsic protein 2; 1) in closed and open conformations and proposed a mechanism for how this gating can be achieved. To study the effect of mercury on SoPIP2; 1 we solved the structure of the SoPIP2;1-mercury complex and characterized the water transport ability using proteoliposomes. The structure revealed mercury binding to three out of four cysteine residues. In contrast to what is normally seen for AQPs, mercury increased the water transport rate of SoPIP2; 1, an effect which could not be attributed to any of the cysteine residues. This indicates that other factors might influence the effect of mercury on SoPIP2; 1, one of which could be the properties of the lipid bilayer.
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| 9. |
- Frick, Anna, 1982, et al.
(author)
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Structural basis for pH gating of plant aquaporins
- 2013
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In: Febs Letters. - : Wiley. - 0014-5793. ; 587:7, s. 989-993
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Journal article (peer-reviewed)abstract
- Plants have evolved to cope with fluctuations in water supply by gating their water channels known
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| 10. |
- Frick, Anna, 1982, et al.
(author)
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X-ray structure of human aquaporin 2 and its implications for nephrogenic diabetes insipidus and trafficking.
- 2014
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In: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 1091-6490 .- 0027-8424. ; 111:17, s. 6305-10
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Journal article (peer-reviewed)abstract
- Human aquaporin 2 (AQP2) is a water channel found in the kidney collecting duct, where it plays a key role in concentrating urine. Water reabsorption is regulated by AQP2 trafficking between intracellular storage vesicles and the apical membrane. This process is tightly controlled by the pituitary hormone arginine vasopressin and defective trafficking results in nephrogenic diabetes insipidus (NDI). Here we present the X-ray structure of human AQP2 at 2.75 Å resolution. The C terminus of AQP2 displays multiple conformations with the C-terminal α-helix of one protomer interacting with the cytoplasmic surface of a symmetry-related AQP2 molecule, suggesting potential protein-protein interactions involved in cellular sorting of AQP2. Two Cd(2+)-ion binding sites are observed within the AQP2 tetramer, inducing a rearrangement of loop D, which facilitates this interaction. The locations of several NDI-causing mutations can be observed in the AQP2 structure, primarily situated within transmembrane domains and the majority of which cause misfolding and ER retention. These observations provide a framework for understanding why mutations in AQP2 cause NDI as well as structural insights into AQP2 interactions that may govern its trafficking.
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