Browsing by Author "Kent, B"
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- ItemIntrinsically disordered stress protein COR15A resides at the membrane surface during dehydration(Biophysical Society, 2017-08-08) Bremer, A; Kent, B; Hauß, T; Thalhammer, A; Yepuri, NR; Darwish, TA; Garvey, CJ; Bryant, G; Hincha, DKPlants from temperate climate zones are able to increase their freezing tolerance during exposure to low, above-zero temperatures in a process termed cold acclimation. During this process, several cold-regulated (COR) proteins are accumulated in the cells. One of them is COR15A, a small, intrinsically disordered protein that contributes to leaf freezing tolerance by stabilizing cellular membranes. The isolated protein folds into amphipathic α-helices in response to increased crowding conditions, such as high concentrations of glycerol. Although there is evidence for direct COR15A-membrane interactions, the orientation and depth of protein insertion were unknown. In addition, although folding due to high osmolyte concentrations had been established, the folding response of the protein under conditions of gradual dehydration had not been investigated. Here we show, using Fourier transform infrared spectroscopy, that COR15A starts to fold into α-helices already under mild dehydration conditions (97% relative humidity (RH), corresponding to freezing at −3°C) and that folding gradually increases with decreasing RH. Neutron diffraction experiments at 97 and 75% RH established that the presence of COR15A had no significant influence on the structure of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membranes. However, using deuterated POPC we could clearly establish that COR15A interacts with the membranes and penetrates below the headgroup region into the upper part of the fatty acyl chain region. This localization is in agreement with our hypothesis that COR15A-membrane interaction is at least, in part, driven by a hydrophobic interaction between the lipids and the hydrophobic face of the amphipathic protein α-helix. © 2017 Biophysical Society - Open access
- ItemInverse hexagonal - inverse ribbon - lamellar gel phase transition sequence in low hydration DOPC:DOPE phospholipid mixtures(Elsevier, 2009-01) Kent, B; Garvey, CJ; Cookson, DJ; Bryant, GThe inverse hexagonal to inverse ribbon phase transition in a mixed phosphatidylcholine-phosphatidylethanolamine system at low hydration is studied using small and wide angle X-ray scattering. It is found that the structural parameters of the inverse hexagonal phase are independent of temperature. By contrast the length of each ribbon of the inverse ribbon phase increases continuously with decreasing temperature over a range of 50 C. At low temperatures the inverse ribbon phase is observed to have a transition to a gel lamellar phase, with no intermediate fluid lamellar phase. This phase transition is confirmed by differential scanning calorimetry. © 2008, Elsevier Ltd.
- ItemMeasurement of glucose exclusion from the fully hydrated DOPE inverse hexagonal phase(Royal Society of Chemistry, 2010-03-21) Kent, B; Garvey, CJ; Lenné, T; Porcar, L; Garamus, VM; Bryant, GThe degree of exclusion of glucose from the inverse hexagonal HII phase of fully hydrated DOPE is determined using contrast variation small angle neutron scattering and small angle X-ray scattering. The presence of glucose is found to favour the formation of the non-lamellar HII phase over the fluid lamellar phase, over a wide range of temperatures, while having no significant effect on the structure of the HII phase. Glucose is preferentially excluded from the lipid–water interface resulting in a glucose concentration in the HII phase of less than half that in the coexisting aqueous phase. The degree of exclusion is quantified and the results are consistent with a hydration layer of pure water adjacent to the lipid head groups from which glucose is excluded. The osmotic gradient created by the difference in glucose concentration is determined and the influence of glucose on the phase behaviour of non-lamellar phase forming lipid systems is discussed. © 2010, Royal Society of Chemistry
- ItemPhospholipid membrane protection by sugar molecules during dehydration-insights into molecular mechanisms using scattering techniques(MDPI AG, 2013-04-01) Garvey, CJ; Lenné, T; Koster, KL; Kent, B; Bryant, GScattering techniques have played a key role in our understanding of the structure and function of phospholipid membranes. These techniques have been applied widely to study how different molecules (e. g., cholesterol) can affect phospholipid membrane structure. However, there has been much less attention paid to the effects of molecules that remain in the aqueous phase. One important example is the role played by small solutes, particularly sugars, in protecting phospholipid membranes during drying or slow freezing. In this paper, we present new results and a general methodology, which illustrate how contrast variation small angle neutron scattering (SANS) and synchrotron-based X-ray scattering (small angle (SAXS) and wide angle (WAXS)) can be used to quantitatively understand the interactions between solutes and phospholipids. Specifically, we show the assignment of lipid phases with synchrotron SAXS and explain how SANS reveals the exclusion of sugars from the aqueous region in the particular example of hexagonal II phases formed by phospholipids.© 2011, MDPI Publishing. © 2013, MDPI Publishing
- ItemSmall molecule interactions with lipid bilayers by neutron diffraction(International Conference on Neutron Scattering, 2017-07-12) Garvey, CJ; Kent, B; Hauß, T; Georgii, R; Demé, B; Cristiglio, V; Darwish, TA; Wu, CM; Mancera, RL; Bryant, GWhile well established for studying the internal structure of bilayers and orientation of peptides and proteins with respect to bilayers, neutron lamellar diffraction is also a powerful tool for studying the average locus of solubilisation of small molecules in stacks of lipid bilayers. The approach has provided unique insights into important issues in anhydrobiology and cryobiology, where previously only molecular dynamics simulations (MD) had been able to provide molecular scale insights. Currently we use the methodology to understand the interaction of cryo-protectants with bilayers, with the aim of providing important validation of MD parameters to further enhance the utility of the method. Here we discuss the experimental approach, both in terms of the use of contrast variation and the use of selective deuteration to simplify the extraction of scattering length density profiles within the bilayer.
- ItemStudy of the effect of Penetratin on the gyroid to diamond phase transition in Myverol(Australian Institute of Physics, 2012-02-01) Goder, JND; Rafi, NAM; Bryant, G; Hunt, T; Kent, B; Garvey, CJCell-penetrating peptides (CPPs), such as penetratin, have aroused a lot of interest in both the academic and applied research areas for their ability to penetrate cell membranes [1]. Even though there has been much work done, mostly on bilayer membranes, we still do not fully understand the mechanisms involved during this phenomenon. In our investigation, we are going to focus on the phase transition of Myverol/Saline system from Gyroid to Diamond cubic phase in the presence of penetratin, to get an understanding of how CPPs work in a more complex system. We use a Myverol/Saline system that exhibits these types of phase behaviors at specific compositions [2]. We plan to use a variety of Small Angle Scattering techniques in an attempt to elucidate the transitional phase behavior of the Myverol/Saline system with varying concentrations of penetratin. From this we hope to develop a model system for investigating the role of a simple peptide in changing the packing of amphiphilic molecules. In this study, we are making the use of 2 complementary techniques, Small Angle X-ray Scattering (SAXS) and small angle neutron scattering (SANS), to investigate the Myverol/Saline system as functions of both temperatures and peptide composition. We also hope to carry out DSC measurements in parallel with the SAXS. We will report on preliminary SAXS experiments carried out at the Australian Synchrotron.