| 1. |
- Björkelund, Hanna, et al.
(författare)
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Gefitinib Induces Epidermal Growth Factor Receptor Dimers Which Alters the Interaction Characteristics with (125)I-EGF
- 2011
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:9, s. e24739-
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Tidskriftsartikel (refereegranskat)abstract
- The tyrosine kinase inhibitor gefitinib inhibits growth in some tumor types by targeting the epidermal growth factor receptor (EGFR). Previous studies show that the affinity of the EGF-EGFR interaction varies between hosting cell line, and that gefitinib increases the affinity for some cell lines. In this paper, we investigate possible mechanisms behind these observations. Real-time interaction analysis in LigandTracer (R) Grey revealed that the HER2 dimerization preventing antibody pertuzumab clearly modified the binding of (125)I-EGF to EGFR on HER2 overexpressing SKOV3 cells in the presence of gefitinib. Pertuzumab did not affect the binding on A431 cells, which express low levels of HER2. Cross-linking measurements showed that gefitinib increased the amount of EGFR dimers 3.0-3.8 times in A431 cells in the absence of EGF. In EGF stimulated SKOV3 cells the amount of EGFR dimers increased 1.8-2.2 times by gefitinib, but this effect was cancelled by pertuzumab. Gefitinib treatment did not alter the number of EGFR or HER2 expressed in tumor cell lines A431, U343, SKOV3 and SKBR3. Real-time binding traces were further analyzed in a novel tool, Interaction Map, which deciphered the different components of the measured interaction and supports EGF binding to multiple binding sites. EGFR and HER2 expression affect the levels of EGFR monomers, homodimers and heterodimers and EGF binds to the various monomeric/dimeric forms of EGFR with unique binding properties. Taken together, we conclude that dimerization explains the varying affinity of EGF - EGFR in different cells, and we propose that gefitinib induces EGFR dimmers, which alters the interaction characteristics with (125)I-EGF.
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| 2. |
- Björkelund, Hanna, 1983-, et al.
(författare)
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Resolving the EGF-EGFR interaction characteristics through a multiple-temperature, multiple-inhibitor, real-time interaction analysis approach
- 2013
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Ingår i: Molecular and Clinical Oncology. - : Spandidos Publications. - 2049-9469 .- 2049-9450. ; 1:2, s. 343-352
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Tidskriftsartikel (refereegranskat)abstract
- Overexpression and aberrant activity of the epidermal growth factor (EGF) have been observed in various cancer types, rendering it an important target in oncology research. The interaction between EGF and its receptor (EGFR), as well as subsequent internalization, is complex and may be affected by various factors including tyrosine kinase inhibitors (TKIs). By combining real‑time binding curves produced in LigandTracer® with internalization assays conducted at different temperatures and with different TKIs, the processes of ligand binding, internalization and excretion was visualized. SKOV3 cells had a slower excretion rate compared to A431 and U343 cells, and the tested TKIs (gefitinib, lapatinib, AG1478 and erlotinib) reduced the degree of internalization. The kinetic analysis of the binding curves further demonstrated TKI‑dependent balances of EGFR monomer and dimer populations, where lapatinib promoted the monomeric form, while the other TKIs induced dimers. The dimer levels were found to be associated with the apparent affinity of the EGF‑EGFR interaction, with EGF binding stronger to EGFR dimers compared to monomers. This study analyzed how real‑time molecular interaction analysis may be utilized in combination with perturbations in order to understand the kinetics of a ligand‑receptor interaction, as well as some of its associated intracellular processes. Our multiple‑temperature and ‑inhibitor assay setup renders it possible to follow the EGFR monomer, dimer and internalized populations in a detailed manner, allowing for a new perspective of the EGFR biology.
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| 3. |
- Bongini, Lorenzo, et al.
(författare)
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Resolving the geometry of biomolecules imaged by cryo electron tomography
- 2007
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Ingår i: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 228:2, s. 174-184
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Tidskriftsartikel (refereegranskat)abstract
- In this paper, we describe two methods for computerized analysis of cryo electron tomography reconstructions of biomolecules. Both methods aim at quantifying the degree of structural flexibility of macromolecules and eventually resolving the inner dynamics through automatized protocols. The first method performs a Brownian dynamics evolution of a simplified molecular model into a fictitious force field generated by the tomograms. This procedure enables us to dock the simplified model into the experimental profiles. The second uses a fuzzy framework to delineate the subparts of the proteins and subsequently determine their interdomain relations. The two methods are discussed and their complementarities highlighted with reference to the case of the immunoglobulin antibody. Both artificial maps, constructed from immunoglobulin G entries in the Protein Data Bank and real tomograms are analyzed. Robustness issues and agreement with previously reported measurements are discussed.
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| 4. |
- Brunzell, Ellen, et al.
(författare)
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Investigation of supramolecular structures in various aqueous solutions of an amyloid forming peptide using small-angle X-ray scattering
- 2024
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Ingår i: Soft Matter. - : Royal Society of Chemistry. - 1744-683X .- 1744-6848. ; 20:10, s. 2272-2279
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Tidskriftsartikel (refereegranskat)abstract
- Aggregation of peptide molecules into amyloid fibrils is a characteristic feature of several degenerative diseases. However, the details behind amyloid-formation, and other self-assembled peptide aggregates, remain poorly understood. In this study, we have used small-angle X-ray scattering (SAXS), static and dynamic light scattering (SLS and DLS) as well as cryogenic transmission electron microscopy (cryo-TEM) to determine the structural geometry of self-assembled peptide aggregates in various dilute aqueous solutions. Pramlintide was used as a model peptide to assess the aggregation behaviour of an amyloid-forming peptide. The effects of adding sodium chloride (NaCl), sodium thiocyanate (NaSCN), and sodium fluoride (NaF) and the co-solvent dimethyl sulfoxide (DMSO) on the aggregation behaviour were studied. Our scattering data analysis demonstrates that small oligomeric fibrils aggregate to form networks of supramolecular assemblies with fractal dimensions. The choice of anion in small amounts of added salt has a significant impact on the size of the fibrils as well as on the fractal dimensions of supramolecular clusters. In DMSO the fractal dimension decreased with increasing DMSO concentration, indicating the formation of a less compact structure of the supramolecular assemblies. The peptide pramlintide forms oligomeric species in solution, which make up a supramolecular network characterised by fractal dimensions. The fractal dimension of the network depends on solvent additive.
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| 5. |
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| 6. |
- Essand, Magnus, et al.
(författare)
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Identification and characterization of a novel splicing variant of vesicular monoamine transporter 1
- 2005
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Ingår i: Journal of Molecular Endocrinology. - : Bioscientifica. - 0952-5041 .- 1479-6813. ; 35:3, s. 489-501
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Tidskriftsartikel (refereegranskat)abstract
- Vesicular monoamine transporter 1 (VMAT1) is an integral protein in the membrane of secretory vesicles of neuroendocrine and endocrine cells that allows the transport of biogenic monoamines, such as serotonin, from the cytoplasm into the secretory vesicles. The full-length VMAT1 transcript is produced from 16 exons. We have identified and characterized an alternatively spliced form of VMAT1 that lacks exon 15, the next to last exon of VMAT1. The new form was therefore denoted VMAT1Delta15. Exon 15 does not contain an even multiple of three nucleotides. As a consequence, there is a shift of reading frame, and exon 16 is translated in an alternative reading frame, yielding a novel protein with a shorter and unrelated C-terminus compared with the native VMAT1 protein. VMAT1 and VMAT1Delta15 mRNAs are simultaneously expressed in normal and neoplastic neuroendocrine cells of the GI tract. However, VMAT1 expression is always higher than VMAT1Delta15 expression. We prove that VMAT1Delta15 is not localized in large, dense core vesicles as the native form but in the endoplasmic reticulum. Furthermore, while VMAT1 can take up serotonin, VMAT1Delta15 cannot, indicating different functions for the two forms of VMAT1.
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| 7. |
- Forooqi Motlaq, Vahid, et al.
(författare)
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Investigation of the enhanced ability of bile salt surfactants to solubilize phospholipid bilayers and form mixed micelles.
- 2021
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Ingår i: Soft Matter. - : Royal Society of Chemistry. - 1744-683X .- 1744-6848. ; 17:33, s. 7769-7780
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Tidskriftsartikel (refereegranskat)abstract
- The self-assembly in mixtures of the anionic bile salt surfactant sodium deoxycholate (NaDC) and the zwitterionic phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) in physiological saline solution has been investigated using light scattering, small-angle X-ray scattering and cryo-transmission electron microscopy. Rather small tri-axial ellipsoidal NaDC-DMPC mixed micelles form at a high content of bile salt in the mixture, which increase in size as an increasing amount of DMPC is incorporated into the micelles. Eventually, the micelles begin to grow substantially in length to form long wormlike micelles. At higher mole fractions of DMPC, the samples become turbid and cryo-TEM measurements reveal the existence of large perforated vesicles (stomatosomes), coexisting with geometrically open disks. To our knowledge, stomatosomes have not been observed before for any bile salt-phospholipid system. Mixed micelles are found to be the sole aggregate structure in a very wide regime of bile salt-phospholipid compositions, i.e. up to about 77 mol% phospholipid in the micelles. This is much higher than the corresponding value of 25 mol% observed for the conventional surfactant hexadecyltrimethylammonium bromide (CTAB) mixed with DMPC in the same solvent. The enhanced ability of bile salt surfactants to solubilize phospholipid bilayers and form mixed micelles is rationalized using bending elasticity theory. From our theoretical analysis, we are able to conclude that amphiphilic molecules rank in the following order of increasing spontaneous curvature: phospholipids < conventional surfactants < bile salts. The bending rigidity of the different amphiphilic molecules increases according to the following sequence: bile salts < conventional surfactants < phospholipids.
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| 8. |
- Forooqi Motlaq, Vahid, et al.
(författare)
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Spontaneous Formation of Ultrasmall Unilamellar Vesicles in Mixtures of an Amphiphilic Drug and a Phospholipid
- 2023
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Ingår i: Langmuir. - : American Chemical Society (ACS). - 0743-7463 .- 1520-5827. ; 39:32, s. 11337-11344
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Tidskriftsartikel (refereegranskat)abstract
- We have observed ultrasmall unilamellar vesicles, withdiametersof less than 20 nm, in mixtures of the tricyclic antidepressant drugamitriptyline hydrochloride (AMT) and the unsaturated zwitterionicphospholipid 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) in physiologicalsaline solution. The size and shape of spontaneously formed self-assembledaggregates have been characterized using complementary techniques,i.e., small-angle neutron and X-ray scattering (SANS and SAXS) andcryo-transmission electron microscopy (cryo-TEM). We observe rodlikemixed micelles in more concentrated samples that grow considerablyin length upon dilution, and a transition from micelles to vesiclesis observed as the concentration approaches the critical micelle concentrationof AMT. Unlike the micelles, the spontaneously formed vesicles decreasein size with each step of dilution, and ultrasmall unilamellar vesicles,with diameters as small as about 15 nm, were observed at the lowestconcentrations. The spontaneously formed ultrasmall unilamellar vesiclesmaintain their size for as long we have investigated them (i.e., severalmonths). To the best of our knowledge, such small vesicles have neverbefore been reported to form spontaneously in a biocompatible phospholipid-basedsystem. Most interestingly, the size of the vesicles was observedto be strongly dependent on the chemical structure of the phospholipid,and in mixtures of AMT and the phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine(DMPC), the vesicles were observed to be considerably larger in size.The self-assembly behavior in the phospholipid-drug surfactantsystem in many ways resembles the formation of equilibrium micellesand vesicles in mixed anionic/cationic surfactant systems.
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| 9. |
- Fouard, Celine, et al.
(författare)
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An Objective Comparison between Gray Weighted Distance Transforms and Weighted Distance Transforms On Curved Spaces
- 2006
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Ingår i: Discrete Geometry for Computer Imagery. - Berlin, Heidelberg : Springer Berlin Heidelberg. - 9783540476511 ; , s. 259-270
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Konferensbidrag (refereegranskat)abstract
- In this paper, we compare two different definitions of distance transform for gray level images: the Gray Weighted Distance Transform (GWDT), and the Weighted Distance Transform On Curved Space (WDTOCS). We show through theoretical and experimental comparisons the differences, the strengths and the weaknesses of these two distances.
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| 10. |
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