Tunable membrane binding of the intrinsically disordered dehydrin Lti30, a cold-induced plant stress protein.

Dehydrins are intrinsically disordered plant proteins whose expression is upregulated under conditions of desiccation and cold stress. Their molecular function in ensuring plant survival is not yet known, but several studies suggest their involvement in membrane stabilization. The dehydrins are characterized by a broad repertoire of conserved and repetitive sequences, out of which the archetypical K-segment has been implicated in membrane binding. To elucidate the molecular mechanism of these K-segments, we examined the interaction between lipid membranes and a dehydrin with a basic functional sequence composition: Lti30, comprising only K-segments. Our results show that Lti30 interacts electrostatically with vesicles of both zwitterionic (phosphatidyl choline) and negatively charged phospholipids (phosphatidyl glycerol, phosphatidyl serine, and phosphatidic acid) with a stronger binding to membranes with high negative surface potential. The membrane interaction lowers the temperature of the main lipid phase transition, consistent with Lti30's proposed role in cold tolerance. Moreover, the membrane binding promotes the assembly of lipid vesicles into large and easily distinguishable aggregates. Using these aggregates as binding markers, we identify three factors that regulate the lipid interaction of Lti30 in vitro: (1) a pH dependent His on/off switch, (2) phosphorylation by protein kinase C, and (3) reversal of membrane binding by proteolytic digest.

Impact of oxidized phospholipids on the structural and dynamic organization of phospholipid membranes: a combined DSC and solid state NMR study.

Membranes undergo severe changes under oxidative stress conditions due to the creation of oxidized phospholipid (OxPL) species, which possess molecular properties quite different from their parental lipid components. These OxPLs play crucial roles in various pathological disorders and their occurrence is involved in the onset of intrinsic apoptosis, a fundamental pathway in programmed mammalian cell death. However, the molecular mechanisms by which these lipids can exert their apoptotic action via their host membranes (e.g., altering membrane protein function) are poorly understood. Therefore, we studied the impact of OxPLs on the organization and biophysical properties of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) based lipid membranes by differential scanning calorimetry (DSC) and solid state nuclear magnetic resonance (NMR) spectroscopy. Incorporation of defined OxPLs with either a carboxyl group (1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine (PazePC)) or aldehyde (1-palmitoyl-(9'oxononanoyl)-sn-glycero-3-phosphocholine (PoxnoPC)) at their truncated sn-2-chain ends enabled us to reveal OxPL species-dependent differences. The calorimetric studies revealed significant effects of OxPLs on the thermotropic phase behavior of DMPC bilayers, especially at elevated levels where PazePC induced more pronounced effects than PoxnoPC. Temperature-dependent changes in the solid state 31P NMR spectra, which provided information of the lipid headgroup region in these mixed membrane systems, reflected this complex phase behavior. In the temperature region between 293 K (onset of the Lalpha-phase) and 298 K, two overlapping NMR spectra were visible which reflect the co-existence of two liquid-crystalline lamellar phases with presumably one reflecting OxPL-poor domains and the other OxPL-rich domains. Deconvolution of the DSC profiles also revealed these two partially overlapping thermal events. In addition, a third thermal, non-NMR-visible, event occurred at low temperatures, which can most likely be associated to a solid-phase mixing/demixing process of the OxPL-containing membranes. The observed phase transitions were moved to higher temperatures in the presence of heavy water due its condensing effect, where additional wideline 2H-NMR studies revealed a complex hydration pattern in the presence of OxPLs.