| 1. |
- Holmström, Anna, et al.
(författare)
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LcrV is a channel size-determining component of the Yop effector translocon of Yersinia
- 2001
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Ingår i: Molecular Microbiology. - : Blackwell Publishing. - 0950-382X .- 1365-2958. ; 39:3, s. 620-632
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Tidskriftsartikel (refereegranskat)abstract
- Delivery of Yop effector proteins by pathogenic Yersinia across the eukaryotic cell membrane requires LcrV, YopB and YopD. These proteins were also required for channel formation in infected erythrocytes and, using different osmolytes, the contact‐dependent haemolysis assay was used to study channel size. Channels associated with LcrV were around 3 nm, whereas the homologous PcrV protein of Pseudomonas aeruginosa induced channels of around 2 nm in diameter. In lipid bilayer membranes, purified LcrV and PcrV induced a stepwise conductance increase of 3 nS and 1 nS, respectively, in 1 M KCl. The regions important for channel size were localized to amino acids 127–195 of LcrV and to amino acids 106–173 of PcrV. The size of the channel correlated with the ability to translocate Yop effectors into host cells. We suggest that LcrV is a size‐determining structural component of the Yop translocon.
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| 2. |
- Bárcena-Uribarri, Iván, et al.
(författare)
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P66 porins are present in both Lyme disease and relapsing fever spirochetes : a comparison of the biophysical properties of P66 porins from six Borrelia species
- 2010
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Ingår i: Biochimica et Biophysica Acta - Biomembranes. - : Elsevier. - 0005-2736 .- 1879-2642. ; 1798:6, s. 1197-1203
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Tidskriftsartikel (refereegranskat)abstract
- The genus Borrelia is the cause of the two human diseases: Lyme disease (LD) and relapsing fever (RF). BothLD and RF Borrelia species are obligate parasites and are dependent on nutrients provided by their hosts. Thefirst step of nutrient uptake across the outer membrane of these Gram-negative bacteria is accomplished bywater-filled channels, so-called porins. The knowledge of the porin composition in the outer membranes ofthe different pathogenic Borrelia species is limited. Only one porin has been described in relapsing feverspirochetes to date, whereas four porins are known to be present in Lyme disease agents. From these, theBorrelia burgdorferi outer membrane channel P66 is known to act as an adhesin and was well studied as aporin. To investigate if P66 porins are expressed and similarly capable of pore formation in other Borreliacausing Lyme disease or relapsing fever three LD species (B. burgdorferi, B. afzelii, B. garinii) and three RFspecies (B. duttonii, B. recurrentis and B. hermsii) were investigated for outer membrane proteins homologousto P66. A search in current published RF genomes, comprising the ones of B. duttonii, B. recurrentis and B.hermsii, indicated that they all contained P66 homologues. The P66 homologues of the six Borrelia specieswere purified to homogeneity and their pore-forming abilities as well as the biophysical properties of thepores were analyzed using the black lipid bilayer assay.
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| 3. |
- Bárcena-Uribarri, Iván, et al.
(författare)
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Study of the protein complex, pore diameter, and pore-forming activity of the Borrelia burgdorferi P13 porin
- 2014
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Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 289:27, s. 18614-18624
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Tidskriftsartikel (refereegranskat)abstract
- P13 is one of the major outer membrane proteins of Borrelia burgdorferi. Previous studies described P13 as a porin. In the present study some structure and function aspects of P13 were studied. P13 showed according to lipid bilayer studies a channel-forming activity of 0.6 nanosiemens in 1 M KCl. Single channel and selectivity measurements demonstrated that P13 had no preference for either cations or anions and showed no voltage-gating up to +/-100 mV. Blue native polyacrylamide gel electrophoresis was used to isolate and characterize the P13 protein complex in its native state. The complex had a high molecular mass of about 300 kDa and was only composed of P13 monomers. The channel size was investigated using non-electrolytes revealing an apparent diameter of about 1.4 nm with a 400-Da molecular mass cut-off. Multichannel titrations with different substrates reinforced the idea that P13 forms a general diffusion channel. The identity of P13 within the complex was confirmed by second dimension SDS-PAGE, Western blotting, mass spectrometry, and the use of a p13 deletion mutant strain. The results suggested that P13 is the protein responsible for the 0.6-nanosiemens pore-forming activity in the outer membrane of B. burgdorferi.
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| 4. |
- Barcena-Uribarri, Ivan, et al.
(författare)
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Use of Nonelectrolytes Reveals the Channel Size and Oligomeric Constitution of the Borrelia burgdorferi P66 Porin
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science. - 1932-6203. ; 8:11, s. e78272-
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Tidskriftsartikel (refereegranskat)abstract
- In the Lyme disease spirochete Borrelia burgdorferi, the outer membrane protein P66 is capable of pore formation with an atypical high single-channel conductance of 11 nS in 1 M KCl, which suggested that it could have a larger diameter than 'normal' Gram-negative bacterial porins. We studied the diameter of the P66 channel by analyzing its single-channel conductance in black lipid bilayers in the presence of different nonelectrolytes with known hydrodynamic radii. We calculated the filling of the channel with these nonelectrolytes and the results suggested that nonelectrolytes (NEs) with hydrodynamic radii of 0.34 nm or smaller pass through the pore, whereas neutral molecules with greater radii only partially filled the channel or were not able to enter it at all. The diameter of the entrance of the P66 channel was determined to be <= 1.9 nm and the channel has a central constriction of about 0.8 nm. The size of the channel appeared to be symmetrical as judged from one-sidedness of addition of NEs. Furthermore, the P66-induced membrane conductance could be blocked by 80-90% by the addition of the nonelectrolytes PEG 400, PEG 600 and maltohexaose to the aqueous phase in the low millimolar range. The analysis of the power density spectra of ion current through P66 after blockage with these NEs revealed no chemical reaction responsible for channel block. Interestingly, the blockage of the single-channel conductance of P66 by these NEs occurred in about eight subconductance states, indicating that the P66 channel could be an oligomer of about eight individual channels. The organization of P66 as a possible octamer was confirmed by Blue Native PAGE and immunoblot analysis, which both demonstrated that P66 forms a complex with a mass of approximately 460 kDa. Two dimension SDS PAGE revealed that P66 is the only polypeptide in the complex.
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| 5. |
- Bárcena-Uribarri, Iván, et al.
(författare)
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Use of nonelectrolytes reveals the channel size and oligomeric constitution of the Borrelia burgdorferi P66 porin
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The outer membrane protein P66 of the Lyme disease spirochete Borrelia burgdorferi is capable of pore formation with an atypical high single-channel conductance of 11 nS in 1 M KCl. We studied in a non-theoretical manner the diameter of the P66 channel by analyzing its single-channel conductance in black lipid bilayers in the presence of different nonelectrolytes with known hydrodynamic radii. Furthermore, we calculated the filling of the channel with these nonelectrolytes and the results revealed that nonelectrolytes with hydrodynamic radii of 0.34 nm or smaller pass through the pore, whereas neutral molecules with greater radii only partially filled the channel or were not able to enter it at all. Thus, the diameter of the P66 entrance was determined to be ≤ 1.9 nm with a constriction site diameter of about 0.7 nm. Furthermore, the P66-induced membrane conductance could be blocked by 80-90% after addition of the nonelectrolytes PEG 400, PEG 600 and maltohexaose in the low millimolar range. Interestingly, the analysis of the power density spectra of P66 after blockage with nonelectrolytes revealed no chemical interaction responsible for channel block. The blockage of one P66 single-channel conductance unit of 11 nS occurred by seven subconducting states, thus indicating a heptameric organization of the P66 oligomer. This organization of P66 as a heptamer was confirmed by Blue Native PAGE and immunoblot analysis, which demonstrated that P66 forms a complex with a mass of approximately 460 kDa.
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| 6. |
- Bunikis, Ignas, 1981-, et al.
(författare)
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An RND-type efflux system in Borrelia burgdorferi is involved in virulence and resistance to antimicrobial compounds
- 2008
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Ingår i: PLoS Pathogenicity. - 1553-7374. ; 4:2, s. e1000009-
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Tidskriftsartikel (refereegranskat)abstract
- Borrelia burgdorferi is remarkable for its ability to thrive in widely different environments due to its ability to infect various organisms. In comparison to enteric Gram-negative bacteria, these spirochetes have only a few transmembrane proteins some of which are thought to play a role in solute and nutrient uptake and excretion of toxic substances. Here, we have identified an outer membrane protein, BesC, which is part of a putative export system comprising the components BesA, BesB and BesC. We show that BesC, a TolC homolog, forms channels in planar lipid bilayers and is involved in antibiotic resistance. A besC knockout was unable to establish infection in mice, signifying the importance of this outer membrane channel in the mammalian host. The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure. We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB. We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.
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| 7. |
- Delrez, L., et al.
(författare)
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Refining the properties of the TOI-178 system with CHEOPS and TESS
- 2023
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Ingår i: Astronomy and Astrophysics. - 0004-6361 .- 1432-0746. ; 678
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Tidskriftsartikel (refereegranskat)abstract
- Context. The TOI-178 system consists of a nearby late K-dwarf transited by six planets in the super-Earth to mini-Neptune regime, with radii ranging from -1.1 to 2.9 R and orbital periods between 1.9 and 20.7 days. All planets but the innermost one form a chain of Laplace resonances. Mass estimates derived from a preliminary radial velocity (RV) dataset suggest that the planetary densities do not decrease in a monotonic way with the orbital distance to the star, contrary to what one would expect based on simple formation and evolution models. Aims. To improve the characterisation of this key system and prepare for future studies (in particular with JWST), we performed a detailed photometric study based on 40 new CHEOPS visits, one new TESS sector, and previously published CHEOPS, TESS, and NGTS data. Methods. First we updated the parameters of the host star using the new parallax from Gaia EDR3. We then performed a global analysis of the 100 transits contained in our data to refine the physical and orbital parameters of the six planets and study their transit timing variations (TTVs). We also used our extensive dataset to place constraints on the radii and orbital periods of potential additional transiting planets in the system. Results. Our analysis significantly refines the transit parameters of the six planets, most notably their radii, for which we now obtain relative precisions of -3%, with the exception of the smallest planet, b, for which the precision is 5.1%. Combined with the RV mass estimates, the measured TTVs allow us to constrain the eccentricities of planets c to g, which are found to be all below 0.02, as expected from stability requirements. Taken alone, the TTVs also suggest a higher mass for planet d than that estimated from the RVs, which had been found to yield a surprisingly low density for this planet. However, the masses derived from the current TTV dataset are very prior-dependent, and further observations, over a longer temporal baseline, are needed to deepen our understanding of this iconic planetary system.
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| 8. |
- Ehrenreich, D., et al.
(författare)
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A full transit of v 2 Lupi d and the search for an exomoon in its Hill sphere with CHEOPS
- 2023
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Ingår i: Astronomy and Astrophysics. - : EDP Sciences. - 0004-6361 .- 1432-0746. ; 671
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Tidskriftsartikel (refereegranskat)abstract
- The planetary system around the naked-eye star v2 Lupi (HD 136352; TOI-2011) is composed of three exoplanets with masses of 4.7, 11.2, and 8.6 Earth masses (M⊕). The TESS and CHEOPS missions revealed that all three planets are transiting and have radii straddling the radius gap separating volatile-rich and volatile-poor super-earths. Only a partial transit of planet d had been covered so we re-observed an inferior conjunction of the long-period 8.6 M⊕ exoplanet v2 Lup d with the CHEOPS space telescope. We confirmed its transiting nature by covering its whole 9.1 h transit for the first time. We refined the planet transit ephemeris to P = 107.13610.0022+0.0019 days and Tc = 2459009.77590.0096+0.0101 BJDTDB, improving by ~40 times on the previously reported transit timing uncertainty. This refined ephemeris will enable further follow-up of this outstanding long-period transiting planet to search for atmospheric signatures or explore the planet s Hill sphere in search for an exomoon. In fact, the CHEOPS observations also cover the transit of a large fraction of the planet s Hill sphere, which is as large as the Earth s, opening the tantalising possibility of catching transiting exomoons. We conducted a search for exomoon signals in this single-epoch light curve but found no conclusive photometric signature of additional transiting bodies larger than Mars. Yet, only a sustained follow-up of v2 Lup d transits will warrant a comprehensive search for a moon around this outstanding exoplanet.
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| 9. |
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| 10. |
- Hoyer, S., et al.
(författare)
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Characterization of the HD 108236 system with CHEOPS and TESS Confirmation of a fifth transiting planet
- 2022
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Ingår i: Astronomy and Astrophysics. - : EDP Sciences. - 0004-6361 .- 1432-0746. ; 668
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Tidskriftsartikel (refereegranskat)abstract
- Context. The HD 108236 system was first announced with the detection of four small planets based on TESS data. Shortly after, the transit of an additional planet with a period of 29.54 d was serendipitously detected by CHEOPS. In this way, HD 108236 (V = 9.2) became one of the brightest stars known to host five small transiting planets (Rp < 3 Ro˙). Aims. We characterize the planetary system by using all the data available from CHEOPS and TESS space missions. We use the flexible pointing capabilities of CHEOPS to follow up the transits of all the planets in the system, including the fifth transiting body. Methods. After updating the host star parameters by using the results from Gaia eDR3, we analyzed 16 and 43 transits observed by CHEOPS and TESS, respectively, to derive the planets' physical and orbital parameters. We carried out a timing analysis of the transits of each of the planets of HD 108236 to search for the presence of transit timing variations. Results. We derived improved values for the radius and mass of the host star (R∗ = 0.876 ± 0.007 R0 and M∗ = 0.867-0.046+0.047M). We confirm the presence of the fifth transiting planet f in a 29.54 d orbit. Thus, the HD 108236 system consists of five planets of Rb = 1.587±0.028, Rc = 2.122±0.025, Rd = 2.629 ± 0.031, Re = 3.008 ± 0.032, and Rf = 1.89 ± 0.04 [Ro˙]. We refine the transit ephemeris for each planet and find no significant transit timing variations for planets c, d, and e. For planets b and f, instead, we measure significant deviations on their transit times (up to 22 and 28 min, respectively) with a non-negligible dispersion of 9.6 and 12.6 min in their time residuals. Conclusions. We confirm the presence of planet f and find no significant evidence for a potential transiting planet in a 10.9 d orbital period, as previously suggested. Further monitoring of the transits, particularly for planets b and f, would confirm the presence of the observed transit time variations. HD 108236 thus becomes a key multi-planetary system for the study of formation and evolution processes. The reported precise results on the planetary radii - together with a profuse RV monitoring - will allow for an accurate characterization of the internal structure of these planets.
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