Conformational Ensembles from Experimental Data and Computer Simulations

Conformational Ensembles from Experimental Data and Computer Simulations

Poster Abstracts

98-POS Board 18 Partial Folding of Intrinsically Disordered Plant LEA Proteins is Required for Membrane Binding and Stabilization Anja Thalhammer 1 , Anne Bremer 2 , Carlos Navarro-Retamal 3 , Gary Bryant 4 , Wendy González 3 , Dirk K. Hincha 2 . 1 University of Potsdam, Potsdam, Germany, 2 Max-Planck Institute of Molecular Plant Physiology, Potsdam, Germany, 3 Universidad de Talca, Talca, Chile, 4 RMIT University, Melbourne, Australia. Late embryogenesis abundant (LEA) proteins accumulate in seeds and vegetative plant tissues, especially after exposure to abiotic stresses and in desiccation tolerant bacteria and invertebrates. Their expression is directly linked to cellular dehydration as arising during freezing or desiccation. Most LEA proteins are intrinsically disordered under fully hydrated conditions and fold during drying. We focus on two cold-induced Arabidopsis thaliana LEA proteins, COR15A and COR15B. Functionally redundant, COR15A and COR15B stabilize membranes during freezing in vitro and in vivo while they do not stabilize selected enzymes during freezing in vivo . Both proteins are disordered in solution, but fold into amphipathic α-helices in the dry state, as shown by circular dichroism (CD) and fourier-transform infrared (FTIR) spectroscopy and in silico analysis. The unfolding process of both COR15 proteins after transfer to water was modeled by Molecular Dynamics simulations, using homology and threading modelling approaches and showed quantitative agreement with experimental data. In water, unfolding was driven by a break of intramolecular and concomitant formation of protein-water H-bonds. We used glycerol as a low-molecular weight crowding agent to model reduced cellular water availability. Experimentally, we found a concentration dependent gain of α-helical structure in solutions containing glycerol. Unfolding of COR15A and COR15B as assessed by Molecular Dynamics simulations was reduced in glycerol-containing systems, indicating that structural stabilization can be explained by preferential exclusion of glycerol from the protein backbone. FTIR spectroscopy, X-ray diffraction and Molecular Dynamics simulations further revealed that COR15A associates with artificial membranes exclusively in an at least partially folded state. Overall, our findings indicate an initial dehydration-induced folding step is necessary to render the COR15 proteins competent for membrane interaction. A second folding step takes place during membrane association.

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