Over the past half century, as molecular biology and biochemistry have developed to inform our understanding of disease, and as this understanding has driven the search for treatments in biotechnology and molecular medicine, the dominant theoretical scaffold for interpreting molecular behavior has been the structure-function relationship. Our understanding of molecular biology's structure-function relationships is largely limited, however, to the molecules of the central dogma: proteins and oligonucleotides. The vast variety of molecular structure outside of the central dogma, particularly among lipids and carbohydrates, suggests that these, too, can be understood in terms of structure driving function. In particular, there has been intense interest in the functional role that lipids might play in the plasma membrane. This project deploys a new class of synthetic lipid bilayers to begin drawing connections between the structure of lipid molecules-both in terms of individual molecular structure and supermolecular organization-and the function of the plasma membrane. The research tools developed and deployed here, called asymmetric giant unilamellar vesicles (AGUVs), are designed to uniquely mimic the cell membrane, capturing properties such as compositional asymmetry and molecular crowding better than other existing synthetic lipid bilayers. One of the many questions that AGUVs can help answer involves passive transport across the cell membrane. Passive transport is an important route for drug delivery and passage of environmental toxins into cells. Recent results show that the mechanism of this transport is complex, and highly dependent on lipid behavior. This project deploys an AGUV-based technique for systematically measuring the dynamics of solute molecules interacting with and penetrating lipid bilayers, yielding richer mechanistic data than other approaches are capable of delivering. AGUVs can also be used to study the mechanical properties of the cell membrane. These properties- particularly resistance to bending-control protein function and are important in a range of physiological processes. While synthetic lipid bilayers have been used to probe these properties, little is known about the effects of bilayer asymmetry on them. This project uses AGUVs to discover these effects. Lipid interactions with integral membrane proteins are likely a major mode by which lipids influence cell behavior. Very little is known, however, about the origins or controlling parameters of these interactions. This project begins to untangle this problem in AGUVs by using fluorescence microscopy to probe how peptides modeling the transmembrane regions of various proteins associate with segregated lipid domains. Finally, AGUVs can facilitate the systematic study of the effects of macromolecular crowding in the cell interior. The microfluidic technique by which AGUVs are formed allows for the inclusion of arbitrary molecules within them, leading to unique molecularly crowded structures.

Public Health Relevance

Cells are surrounded by membranes that consist of two layers of lipid molecules, and the chemical composition is different in these two layers. In this project, a new type of artificial cell membrane that is uniquely capable of mimicking this asymmetry is deployed to study a range of important biological processes, including drug transport into cells, mechanical deformation of the cell membrane, and interactions between lipid molecules and receptor proteins involved in cancer treatment and diabetes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM093279-04
Application #
8534183
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Chin, Jean
Project Start
2010-09-30
Project End
2015-07-31
Budget Start
2013-08-01
Budget End
2014-07-31
Support Year
4
Fiscal Year
2013
Total Cost
$281,543
Indirect Cost
$100,026
Name
University of Southern California
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
072933393
City
Los Angeles
State
CA
Country
United States
Zip Code
90089
Mora, Nestor Lopez; Gao, Yue; Gutierrez, M Gertrude et al. (2017) Evaluation of dextran(ethylene glycol) hydrogel films for giant unilamellar lipid vesicle production and their application for the encapsulation of polymersomes. Soft Matter 13:5580-5588
Peruzzi, Justin; Gutierrez, M Gertrude; Mansfield, Kylee et al. (2016) Dynamics of Hydrogel-Assisted Giant Unilamellar Vesicle Formation from Unsaturated Lipid Systems. Langmuir 32:12702-12709
Gutierrez, M Gertrude; Jalali-Yazdi, Farzad; Peruzzi, Justin et al. (2016) G Protein-Coupled Receptors Incorporated into Rehydrated Diblock Copolymer Vesicles Retain Functionality. Small 12:5256-5260
Gutierrez, M Gertrude; Mansfield, Kylee S; Malmstadt, Noah (2016) The Functional Activity of the Human Serotonin 5-HT1A Receptor Is Controlled by Lipid Bilayer Composition. Biophys J 110:2486-2495
Bhargava, Krisna C; Thompson, Bryant; Tembhekar, Anoop et al. (2016) Temperature Sensing in Modular Microfluidic Architectures. Micromachines (Basel) 7:
Runas, Kristina A; Acharya, Shiv J; Schmidt, Jacob J et al. (2016) Addition of Cleaved Tail Fragments during Lipid Oxidation Stabilizes Membrane Permeability Behavior. Langmuir 32:779-86
Bhargava, Krisna C; Thompson, Bryant; Iqbal, Danish et al. (2015) Predicting the behavior of microfluidic circuits made from discrete elements. Sci Rep 5:15609
Wu, Shao-Hua; Sankhagowit, Shalene; Biswas, Roshni et al. (2015) Viscoelastic deformation of lipid bilayer vesicles. Soft Matter 11:7385-91
Runas, Kristina A; Malmstadt, Noah (2015) Low levels of lipid oxidation radically increase the passive permeability of lipid bilayers. Soft Matter 11:499-505
Hu, Peichi C; Malmstadt, Noah (2015) Asymmetric giant lipid vesicle fabrication. Methods Mol Biol 1232:79-90

Showing the most recent 10 out of 26 publications