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Investigating Lipid-Protein Interactions in a Complex Biological Membrane Model
Tuesday, March 1, 2016
Protein-Lipid Interactions II 2074-Pos Board B218 Investigating Lipid-Protein Interactions in a Complex Biological Membrane Model Karelia H. Delgado-Magnero, Gurpreet Singh, Valentina Corradi, D. Peter Tieleman. Centre for Molecular Simulation, Biological Sciences, University of Calgary, Calgary, AB, Canada. Biological membranes are crucial as they define essential compartments for cells. Cell membranes are heterogeneous mixtures of membrane proteins and lipids. Their structure and function are fundamentally dependent on lipidlipid and lipid-protein interactions which play a crucial role in regulating cell protein functions and are involved in many diseases when altered. However, the exact properties and the relation among the distinct components that form the biological membrane are not completely understood due the limitations of experimental methods in studying the lipid-protein interactions in living cells. Lately, computer simulations have become a promising tool to understand and clarify experimental results. The goal of our research is to identify specific lipid-protein interactions of biological relevance and work towards large-scale modeling of realistic biological membranes. In this study, we placed ten main classes of eukaryotic membrane proteins with different ratios in a prototypical plasma membrane model, containing various lipid types found in the plasma membranes of eukaryotic cells. The protein-membrane system was prepared using Martini force field. Using molecular dynamic simulations, we intend to investigate how proteins sort lipids, and to identify specific binding partners and lipids of interest for more detailed studies. 2075-Pos Board B219 Activation of Integrin: A Multiscale Computational Study Anirban Polley1, Anand Srivastava2, Gregory A. Voth1. 1 Chemistry, The University of Chicago, Chicago, IL, USA, 2Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India. Despite the importance of integrin proteins in cell signal transduction and force generation, the mechanism of action is not understood at the atomic level. To date, no experimental method has been able to probe the structure of integrin proteins in their open, force-activated state. To gain a better understanding of the transformation of the inactive, closed, state to an active state of the integrin, we are employing advanced computational techniques that will allow us to more quickly sample the conformational change and probe the pathway of the transformation. To make connection with the experimental studies, we have first studied the effect of single and double amino acid substitutions to the wild type integrin. Our simulations reveal certain mutants which destabilize the closed state by increasing the kink angle of the extracellular region and distance between the transmembrane region of the alpha and beta subunits of the integrin. We have also developed a coarse-grained model based on our atomistic simulation data, and are using that model to probe the transition of integrin to the open state. 2076-Pos Board B220 The Interaction of Proteins with Asymmetric Lipid Bilayers Milka Doktorova1,2, Gerald W. Feigenson3, Harel Weinstein4. 1 Tri-Institutional Program in Computational Biology and Medicine, Weill Cornell Medical College, New York, NY, USA, 2Cornell University, Ithaca, NY, USA, 3Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY, USA, 4Department of Physiology and Biophysics, Weill Cornell Medical College, New York, NY, USA. Cell membranes can be characterized by their thermodynamic, structural and mechanical properties, such as phase behavior, thickness and rigidity. Experimental and computational studies of membrane properties and their effects on the function of proteins embedded in the lipid environment have focused largely on symmetric bilayers, i.e., with the same lipid composition in the two leaflets. However, the mammalian plasma membrane (PM) has an asymmetric lipid distribution, with an outer leaflet having a significant fraction of high-melting lipids, and an inner leaflet containing low-melting and negatively charged lipids. This asymmetry is often ignored in both experimental and modeling studies especially given the difficulty of preparing asymmetric liposomes in vitro. However, the different elastic and phase properties of symmetric bilayer models of the two PM leaflets, as well as the energy that cells spend to maintain their PM transverse structure, make it likely that this compositional asymmetry provides unique bilayer properties of functional significance. Our goal is to evaluate this hypothesis and its consequences both in vitro and in silico, with a systematic study of symmetric and asymmetric bilayers composed
419a
of phosphatidylcholine, sphingomyelin, phosphatidylserine and/or phosphatidylethanolamine lipid molecules both in the presence and absence of the transmembrane peptides WALP and Gramicidin. We examine the structural manifestations and mechanisms of interleaflet coupling and their effect on protein-membrane interactions, focusing on changes in bending rigidity, acyl chain order, and surface tension. We then evaluate the implications of these properties for the unique deformation of the asymmetric bilayers around the protein inclusions. Results from computation regarding inter-leaflet coupling are compared to in vitro measurements of the thermodynamics properties of asymmetric giant unilamellar vesicles prepared with a newly developed cyclodextrin-mediated exchange protocol. 2077-Pos Board B221 Conformational Dynamics of Prion Proteins at the Membrane Interface Jesse Woo1, Chad Nieri2, Roger Gonzalez1, Jason C. Bartz3, Patricia Soto4. 1 Biology, Creighton University, Omaha, NE, USA, 2Chemistry, Creighton University, Omaha, NE, USA, 3Medical Microbiology and Immunology, Creighton University, Omaha, NE, USA, 4Physics, Creighton University, Omaha, NE, USA. Prions are infectious agents responsible for transmissible spongiform encephalopathies (TSEs), a class of fatal neurodegenerative diseases in many mammals, including humans. The infectious prion protein, PrPSc, propagates by converting the non-pathological conformation of the prion protein, PrPc, into the misfolded PrPSc. It is unclear how infectious PrPSc initially arose, and the mechanism behind the misfolding, conversion, and replication of PrPSc is still widely debated. Since PrPc is found extracellularly and GPI-anchored to the cell membrane, gaining insight on prion-membrane interactions is required to shed light on these uncertainties. In this presentation, we will discuss our results based on biomolecular modeling techniques to identify favorable modes of interaction of the PrPc molecule with model membranes. 2078-Pos Board B222 Structural Analysis of the Interaction of the Functional Amyloid Forming Protein Orb2A with Lipids Maria A. Conrad-Soria, Silvia A. Cervantes, Alexander S. Falk, Thalia H. Bajakian, Ansgar B. Siemer. Zilkha Neurogenetic Institute, University of Southern California, Los Angeles, CA, USA. Orb2 is a CPEB homologue in fruit flies involved in regulation of mRNA translation at the synapse and is essential for stabilization of long term memory. This function is accomplished by the aggregation of Orb2 into amyloid-like oligomers. There are two isoforms of Orb2: Orb2A and Orb2B, and both are found in the aggregates together. However, while Orb2B is constitutively expressed, Orb2A expression is rare and necessary for Orb2 to form aggregates in vivo. This suggests that Orb2A is a main regulator of initiation of fibril formation. This study focuses on Orb2A lipid interaction, a potential mechanism by which Orb2A initiates aggregation. Orb2A is found enriched in the membrane fraction of the synapse, and sequence analysis reveals the possibility of an amphipathic helix present in the unique N-terminus of Orb2A. The aggregation of several other amyloids, including IAPP, Pmel17, and Ab, has been shown to be regulated by lipids. I hypothesize that such a mechanism could also be utilized by Orb2A to either inhibit or induce amyloid formation. I use circular dichroism, electron paramagnetic resonance, and solid-state NMR of Orb2A deletion mutants to explore the effects of the interaction of Orb2A with lipid bilayers. We show that structural changes do occur upon lipid addition and identify the region involved in these structural changes. 2079-Pos Board B223 Interaction of a Model Amphipathic a-Helix Bundle Protein with an Aqueous-Glycerophospholipid-Oil Interface Mona Sadat Mirheydari1, Edgar E. Kooijman2, Elizabeth K. Mann1. 1 Physics, Kent State University, Kent, OH, USA, 2Biological sciences, Kent State University, Kent, OH, USA. The dynamics and formation of lipid droplets has drawn increasing attention. Lipid droplets not only store but control energy sources. They also play roles in building up new membranes, synthesizing steroid hormones and are involved in lipid signaling. The dynamics of protein binding to lipid droplets can be studied in a model system by first forming a lipid monolayer around an oil, triolein, droplet. Protein binding to the monolayer is studied by the decrease in the oil/ water surface tension. Previous groups concentrated on the lipid POPC, one of the major lipids found on lipid droplets, and attempted to tease out the role of amphipathic protein domains by using different protein constructs. However, many of these proteins also contain a distinct domain in which the amphipathic helices can arrange in a bundle which is soluble in solution. PAT family
Protein-Lipid Interactions II 2074-Pos Board B218 Investigating Lipid-Protein Interactions in a Complex Biological Membrane Model Karelia H. Delgado-Magnero, Gurpreet Singh, Valentina Corradi, D. Peter Tieleman. Centre for Molecular Simulation, Biological Sciences, University of Calgary, Calgary, AB, Canada. Biological membranes are crucial as they define essential compartments for cells. Cell membranes are heterogeneous mixtures of membrane proteins and lipids. Their structure and function are fundamentally dependent on lipidlipid and lipid-protein interactions which play a crucial role in regulating cell protein functions and are involved in many diseases when altered. However, the exact properties and the relation among the distinct components that form the biological membrane are not completely understood due the limitations of experimental methods in studying the lipid-protein interactions in living cells. Lately, computer simulations have become a promising tool to understand and clarify experimental results. The goal of our research is to identify specific lipid-protein interactions of biological relevance and work towards large-scale modeling of realistic biological membranes. In this study, we placed ten main classes of eukaryotic membrane proteins with different ratios in a prototypical plasma membrane model, containing various lipid types found in the plasma membranes of eukaryotic cells. The protein-membrane system was prepared using Martini force field. Using molecular dynamic simulations, we intend to investigate how proteins sort lipids, and to identify specific binding partners and lipids of interest for more detailed studies. 2075-Pos Board B219 Activation of Integrin: A Multiscale Computational Study Anirban Polley1, Anand Srivastava2, Gregory A. Voth1. 1 Chemistry, The University of Chicago, Chicago, IL, USA, 2Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India. Despite the importance of integrin proteins in cell signal transduction and force generation, the mechanism of action is not understood at the atomic level. To date, no experimental method has been able to probe the structure of integrin proteins in their open, force-activated state. To gain a better understanding of the transformation of the inactive, closed, state to an active state of the integrin, we are employing advanced computational techniques that will allow us to more quickly sample the conformational change and probe the pathway of the transformation. To make connection with the experimental studies, we have first studied the effect of single and double amino acid substitutions to the wild type integrin. Our simulations reveal certain mutants which destabilize the closed state by increasing the kink angle of the extracellular region and distance between the transmembrane region of the alpha and beta subunits of the integrin. We have also developed a coarse-grained model based on our atomistic simulation data, and are using that model to probe the transition of integrin to the open state. 2076-Pos Board B220 The Interaction of Proteins with Asymmetric Lipid Bilayers Milka Doktorova1,2, Gerald W. Feigenson3, Harel Weinstein4. 1 Tri-Institutional Program in Computational Biology and Medicine, Weill Cornell Medical College, New York, NY, USA, 2Cornell University, Ithaca, NY, USA, 3Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY, USA, 4Department of Physiology and Biophysics, Weill Cornell Medical College, New York, NY, USA. Cell membranes can be characterized by their thermodynamic, structural and mechanical properties, such as phase behavior, thickness and rigidity. Experimental and computational studies of membrane properties and their effects on the function of proteins embedded in the lipid environment have focused largely on symmetric bilayers, i.e., with the same lipid composition in the two leaflets. However, the mammalian plasma membrane (PM) has an asymmetric lipid distribution, with an outer leaflet having a significant fraction of high-melting lipids, and an inner leaflet containing low-melting and negatively charged lipids. This asymmetry is often ignored in both experimental and modeling studies especially given the difficulty of preparing asymmetric liposomes in vitro. However, the different elastic and phase properties of symmetric bilayer models of the two PM leaflets, as well as the energy that cells spend to maintain their PM transverse structure, make it likely that this compositional asymmetry provides unique bilayer properties of functional significance. Our goal is to evaluate this hypothesis and its consequences both in vitro and in silico, with a systematic study of symmetric and asymmetric bilayers composed
419a
of phosphatidylcholine, sphingomyelin, phosphatidylserine and/or phosphatidylethanolamine lipid molecules both in the presence and absence of the transmembrane peptides WALP and Gramicidin. We examine the structural manifestations and mechanisms of interleaflet coupling and their effect on protein-membrane interactions, focusing on changes in bending rigidity, acyl chain order, and surface tension. We then evaluate the implications of these properties for the unique deformation of the asymmetric bilayers around the protein inclusions. Results from computation regarding inter-leaflet coupling are compared to in vitro measurements of the thermodynamics properties of asymmetric giant unilamellar vesicles prepared with a newly developed cyclodextrin-mediated exchange protocol. 2077-Pos Board B221 Conformational Dynamics of Prion Proteins at the Membrane Interface Jesse Woo1, Chad Nieri2, Roger Gonzalez1, Jason C. Bartz3, Patricia Soto4. 1 Biology, Creighton University, Omaha, NE, USA, 2Chemistry, Creighton University, Omaha, NE, USA, 3Medical Microbiology and Immunology, Creighton University, Omaha, NE, USA, 4Physics, Creighton University, Omaha, NE, USA. Prions are infectious agents responsible for transmissible spongiform encephalopathies (TSEs), a class of fatal neurodegenerative diseases in many mammals, including humans. The infectious prion protein, PrPSc, propagates by converting the non-pathological conformation of the prion protein, PrPc, into the misfolded PrPSc. It is unclear how infectious PrPSc initially arose, and the mechanism behind the misfolding, conversion, and replication of PrPSc is still widely debated. Since PrPc is found extracellularly and GPI-anchored to the cell membrane, gaining insight on prion-membrane interactions is required to shed light on these uncertainties. In this presentation, we will discuss our results based on biomolecular modeling techniques to identify favorable modes of interaction of the PrPc molecule with model membranes. 2078-Pos Board B222 Structural Analysis of the Interaction of the Functional Amyloid Forming Protein Orb2A with Lipids Maria A. Conrad-Soria, Silvia A. Cervantes, Alexander S. Falk, Thalia H. Bajakian, Ansgar B. Siemer. Zilkha Neurogenetic Institute, University of Southern California, Los Angeles, CA, USA. Orb2 is a CPEB homologue in fruit flies involved in regulation of mRNA translation at the synapse and is essential for stabilization of long term memory. This function is accomplished by the aggregation of Orb2 into amyloid-like oligomers. There are two isoforms of Orb2: Orb2A and Orb2B, and both are found in the aggregates together. However, while Orb2B is constitutively expressed, Orb2A expression is rare and necessary for Orb2 to form aggregates in vivo. This suggests that Orb2A is a main regulator of initiation of fibril formation. This study focuses on Orb2A lipid interaction, a potential mechanism by which Orb2A initiates aggregation. Orb2A is found enriched in the membrane fraction of the synapse, and sequence analysis reveals the possibility of an amphipathic helix present in the unique N-terminus of Orb2A. The aggregation of several other amyloids, including IAPP, Pmel17, and Ab, has been shown to be regulated by lipids. I hypothesize that such a mechanism could also be utilized by Orb2A to either inhibit or induce amyloid formation. I use circular dichroism, electron paramagnetic resonance, and solid-state NMR of Orb2A deletion mutants to explore the effects of the interaction of Orb2A with lipid bilayers. We show that structural changes do occur upon lipid addition and identify the region involved in these structural changes. 2079-Pos Board B223 Interaction of a Model Amphipathic a-Helix Bundle Protein with an Aqueous-Glycerophospholipid-Oil Interface Mona Sadat Mirheydari1, Edgar E. Kooijman2, Elizabeth K. Mann1. 1 Physics, Kent State University, Kent, OH, USA, 2Biological sciences, Kent State University, Kent, OH, USA. The dynamics and formation of lipid droplets has drawn increasing attention. Lipid droplets not only store but control energy sources. They also play roles in building up new membranes, synthesizing steroid hormones and are involved in lipid signaling. The dynamics of protein binding to lipid droplets can be studied in a model system by first forming a lipid monolayer around an oil, triolein, droplet. Protein binding to the monolayer is studied by the decrease in the oil/ water surface tension. Previous groups concentrated on the lipid POPC, one of the major lipids found on lipid droplets, and attempted to tease out the role of amphipathic protein domains by using different protein constructs. However, many of these proteins also contain a distinct domain in which the amphipathic helices can arrange in a bundle which is soluble in solution. PAT family