Membrane-active protein interactions with phospholipid bilayers

Khatami, Mohammad Hassan (2016) Membrane-active protein interactions with phospholipid bilayers. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

Membrane-active proteins are a class of proteins that interact with lipid membranes in the body. I study two kinds of membrane-active proteins, antimicrobial peptides (AMPs) and lung surfactant (LS) proteins. In the first part of my PhD project I did computer simulation studies with two AMPs, Gaduscidin-1 and -2 (GAD-1 and GAD-2). These peptides are histidine rich and thus expected to exhibit pH-dependent activity. In this work I have performed molecular dynamics (MD) simulations with the peptides in both histidine-charged and histidine-neutral forms, along with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid molecules, employing GROMACS software and an OPLS-AA force field. My results show a high tendency for pairs of histidines to interact with pore regions in both histidine-charged and histidine-neutral simulations. This work is published in Biophysica et Biochimica Acta (BBA)-Biomembranes (2014). In the second part of my PhD research I performed computational simulations on lung surfactant protein B (SP-B) interacting with lipid bilayer. SP-B is a hydrophobic protein with 79 residues, from the saposin superfamily. Because of the extreme hydrophobicity of SP-B, the experimental structure of the protein is unknown. Thus, I combined the Mini-B (a fragment of SP-B) experimental structure and homology modelling based on proteins in saposin family to construct my initial model of SP-B. I run MD (using OPLS-AA and PACE force fields) and replica-exchange MD (using PACE force field) simulations with GROMACS software. I modelled SP-B in open and bent (V-shaped) structures, placed within or near a POPC lipid bilayer. My results demonstrate energetically feasible structures for SP-B, in which salt bridges Membrane-active proteins are a class of proteins that interact with lipid membranes in the body. I study two kinds of membrane-active proteins, antimicrobial peptides (AMPs) and lung surfactant (LS) proteins. In the first part of my PhD project I did computer simulation studies with two AMPs, Gaduscidin-1 and -2 (GAD-1 and GAD-2). These peptides are histidine rich and thus expected to exhibit pH-dependent activity. In this work I have performed molecular dynamics (MD) simulations with the peptides in both histidine-charged and histidine-neutral forms, along with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid molecules, employing GROMACS software and an OPLS-AA force field. My results show a high tendency for pairs of histidines to interact with pore regions in both histidine-charged and histidine-neutral simulations. This work is published in Biophysica et Biochimica Acta (BBA)-Biomembranes (2014). In the second part of my PhD research I performed computational simulations on lung surfactant protein B (SP-B) interacting with lipid bilayer. SP-B is a hydrophobic protein with 79 residues, from the saposin superfamily. Because of the extreme hydrophobicity of SP-B, the experimental structure of the protein is unknown. Thus, I combined the Mini-B (a fragment of SP-B) experimental structure and homology modelling based on proteins in saposin family to construct my initial model of SP-B. I run MD (using OPLS-AA and PACE force fields) and replica-exchange MD (using PACE force field) simulations with GROMACS software. I modelled SP-B in open and bent (V-shaped) structures, placed within or near a POPC lipid bilayer. My results demonstrate energetically feasible structures for SP-B, in which salt bridges Membrane-active proteins are a class of proteins that interact with lipid membranes in the body. I study two kinds of membrane-active proteins, antimicrobial peptides (AMPs) and lung surfactant (LS) proteins. In the first part of my PhD project I did computer simulation studies with two AMPs, Gaduscidin-1 and -2 (GAD-1 and GAD-2). These peptides are histidine rich and thus expected to exhibit pH-dependent activity. In this work I have performed molecular dynamics (MD) simulations with the peptides in both histidine-charged and histidine-neutral forms, along with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid molecules, employing GROMACS software and an OPLS-AA force field. My results show a high tendency for pairs of histidines to interact with pore regions in both histidine-charged and histidine-neutral simulations. This work is published in Biophysica et Biochimica Acta (BBA)-Biomembranes (2014). In the second part of my PhD research I performed computational simulations on lung surfactant protein B (SP-B) interacting with lipid bilayer. SP-B is a hydrophobic protein with 79 residues, from the saposin superfamily. Because of the extreme hydrophobicity of SP-B, the experimental structure of the protein is unknown. Thus, I combined the Mini-B (a fragment of SP-B) experimental structure and homology modelling based on proteins in saposin family to construct my initial model of SP-B. I run MD (using OPLS-AA and PACE force fields) and replica-exchange MD (using PACE force field) simulations with GROMACS software. I modelled SP-B in open and bent (V-shaped) structures, placed within or near a POPC lipid bilayer. My results demonstrate energetically feasible structures for SP-B, in which salt bridges play a significant role. My simulations provide hypotheses for how SP-B promotes the rearrangement of planar lipid bilayers. Part of this work has been accepted for publication in Biophysica et Biochimica Acta (BBA)-Biomembranes (2016). In the third part of my project I employed solid state nuclear magnetic resonance (NMR) using ²H, ³¹P and ¹⁵N experiments, to study SP-B interacting with mechanically oriented lipid bilayer. Here, I used full-length ¹⁵N-labelled SP-B, which was recombinantly expressed in our lab, to find the orientation of protein with respect to the bilayer. In this part of my thesis, the final goal was to compare the experimental ¹⁵N spectra with the spectra, predicted from the structures we got from computational simulations to help define the protein’s structure. Since, I was not able to gain ¹⁵N NMR signals in my SP-B in lipid bilayer experiments, I did not fulfill the final goal of this part of my project. However, I was able to predict ¹⁵N NMR spectra of my computational SP-B structures. My NMR results indicate that more optimization needs to be done to modify our SP-B preparation protocol to 1) increase the yields of isotope-labelled protein and 2) increase the protein:lipid ratio when refolding into lipids. My simulated ¹⁵N spectra indicate that uniform ¹⁵N-labelling is unlikely to constrain SP-B’s structure and topology very much and it will likely be necessary to use a more specifically labelled sample for these experiments.

Item Type: Thesis (Doctoral (PhD))
URI: http://research.library.mun.ca/id/eprint/12448
Item ID: 12448
Additional Information: Includes bibliographical references (pages 184-188).
Keywords: Membrane-Active Proteins, Molecular Dynamics Simulations, Solid-State NMR, GROMACS, Antimicrobial Peptides, Lung surfactant Proteins, SP-B
Department(s): Science, Faculty of > Physics and Physical Oceanography
Date: September 2016
Date Type: Submission
Library of Congress Subject Heading: Peptide antibiotics--Computer simulations; Pulmonary surfactant--Computer simulations; Membrane lipids--Computer simulations

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