Chemical Engineering Dissertation Defense: Ellie Kamaloo

Friday, December 08, 2017
10:00 am to 12:00 pm

Location:

Floor/Room #: 
126

Supported lipid bilayer interactions with nanoparticles, peptides and polymers

Committee members:

  • Terri A Camesano, Ph.D. Professor, Research Advisor Department of Chemical Engineering Worcester Polytechnic Institute
  • Ramanathan Nagarajan, Ph.D. Research Chemical Engineer Natick Soldier Research, Development & Engineering Center (NSRDEC)
  • Amy M. Peterson, Ph.D. Assistant Professor of Chemical Engineering Worcester Polytechnic Institute
  • Jianyu Liang, Ph.D. Associate Professor of Mechanical Engineering Worcester Polytechnic Institute

 

Abstract

Supported lipid bilayers (SLBs) are one of the most common model membranes used in the field of cell membrane biology because they provide a well-defined model membrane platform for determination of molecular-level interactions between different biomolecules (e.g., proteins, peptides) and lipid membranes. This study demonstrates that QCM-D analysis of SLBs serves as powerful tool to investigate and characterize the mechanisms of interactions between lipid membranes and gold nanoparticles (NPs), environmentally relevant polymers, and disease-inducing peptides.

In this study, we investigated the effects of NP size and NP surface chemistry on the interactions between gold NPs and SLB composed of L-α-phosphatidylcholine (egg PC). In order to understand the effect of NP size, gold NPs with diameters of 2, 5, 10, and 40 nm were tested and compared to each other. NPs were tested in their citric acid–stabilized state as well as in the presence of poly (methacrylic acid) (PMAA), which represents an organic coating that could become associated with NPs in the environment.  The results indicated that when dissolved in water, gold NPs with diameters of 2, 5, 10, and 40 nm did not perturb the membrane, but in the presence of environmentally relevant polymers, the larger nanoparticles were found to disrupt the membrane.

In order to better understand the NP-SLB interactions in an  environmentally relevant conditions, the mass and viscoelasticity of the SLB was characterized in the presence of four different natural polymers, also known as natural organic materials (NOMs):  fulvic and humic acids extracted from Suwannee River (SRFA and SRHA), which had relatively lower molecular weights; a commercial humic acid (HA); and humic acid extracted from Elliott soil (ESHA), which had a higher molecular weight. The results showed that NOMs with lower molecular weights adsorbed to the bilayer, while higher molecular weight components did not induce any changes to the bilayers. In addition, the NPs in SRFA and SRHA increased the mass of the bilayer by 20 to 30 ng, while the NPs in HA and ESHA changed the mass of the bilayer by <10 ng. It was concluded that the presence of humic substances, as well as their physical and chemical properties, exerts a direct impact on the interactions between cell membranes and NPs.

In order to elucidate the effect of surface chemistry, 10-nm gold NPs with various functionalizations (i.e., anionic, cationic, and nonionic ligands) were tested.  In these cases, regardless of the type of NP functionalization, no substantial bilayer mass changes were observed. This suggests that the charge and chemistry of the ligands had a minor effect on NP–membrane interactions. Furthermore, in both the control and humic acid experiments, there were small dissipation changes (less than 1 unit) indicating that the overall membrane structure was not perturbed.

In the second part of the project, we focused on SLB interactions with Amyloid β peptide, which is the peptide associated with Alzheimer’s disease (AD). Adsorption of Aβ peptide to cell membrane is known to take place on the so-called “lipid rafts”, which are membrane microdomains enriched with cholesterol, sphingomyelin, and ganglioside. Formation of SLB containing the lipid raft is challenging because it often leads to adsorption of intact vesicles on the substrate without formation of the bilayer. Therefore, the first step of this study was to develop a robust method to form the raft-containing SLB.

In this study, the formation of lipid bilayer composed of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS), cholesterol (Chol), sphingomyelin (SM), and ganglioside (GM) was investigated using QCM-D. Different parameters, such as pH, temperature, osmotic pressure, and vesicle size, were optimized to induce the vesicle rupture. A key parameter in forming the bilayer was found to be applying osmotic pressure to the vesicles by having the vesicles’ exterior concentration of NaCl higher than the interior concentration. Here, the results are discussed based on the mechanisms of vesicle–vesicle and vesicle–substrate interactions.

After developing a robust method for the formation of SLB with lipid rafts, we used that as a template to characterize the mechanism of SLB-Aβ interactions leading to membrane disruption. The mechanism of Aβ toxicity leading to AD has not been fully discovered yet due to the complexity of the process, which includes several steps: Aβ peptide adsorption on the membrane, the conformational change from disordered in solution to a membrane-bound α-helix structure, and then formation of β-sheet aggregates that serve as fibrillation seeds. In this study, we showed that the QCM-D technique is a promising tool to conduct systematic studies on the mechanism of interactions between Aβ peptide and lipid membranes. To our knowledge, this was the first time QCM-D was utilized for characterization of Aβ fibrillation from monomer states through the formation of mature fibrils. The data indicated that peptide–membrane interactions follow a two-step kinetic pathway starting with the adsorption of small (low-n) oligomers until they cover all the adsorption sites on the surface. In the second step, the membrane structure is destabilized as a result of interaction with oligomers, which leads to lipid loss from the surface. Consistency of the results with data obtained via other techniques substantiates QCM-D technique as a robust approach to answer the remaining unanswered questions in the field of Alzheimer’s disease.