Over 30% of proteins are bound to cell membranes, which makes them difficult to isolate in a soluble and active form. We propose to implement recently developed nanolipoprotein particles (NLPs) to allow the separation of membrane-bound proteins in an active form. NLPs are discoidal nanoparticles formed when an apolipoprotein and a population of phospholipids self-assemble into a lipid bilayer in an aqueous environment that can solubilize active membrane-bound proteins. In particular, we will focus on characterization of tyrosine kinase receptors, which mediate the spread of many cancers and are targets of several chemotherapeutics. The ability to study these receptors outside the cell environment will enable structural and mechanistic studies for improved understanding of carcinogenesis and other diseases. Given that membrane-associated proteins account for the majority of drug targets, it is important to develop novel technologies to gain access to this important class of proteins. This main goal is to develop generally applicable methods to produce, isolate and characterize the human type I tyrosine kinase receptors ErbB1, ErbB2, ErbB3 and ErbB4. The epidermal growth factor families of receptors are known to be important in cell signaling and carcinogenesis. These proteins are challenging to study because of their insolubility and tendency to aggregate in aqueous solutions. This proposal is focused on developing a new biotechnology application leading to formation of NLPs capable of solubilizing these and other membrane-bound proteins. The NLPs present a distinct advantage over currently used model membranes in terms of particle size monodispersity and solubility. This proposal will be carried out by synthesizing homo- and heterodimers of ErbB receptors in NLPs, characterizing them for structure and function and performing library screening in order to identify novel high-affinity peptides and small molecules that can serve as lead compounds for further development as drugs.

Public Health Relevance

We propose to apply nanotechnology to the problem of studying membrane-bound proteins, which are difficult to study owing to poor aqueous solubility and protein denaturation. The epidermal growth factor (ErbB) family of membrane-bound receptor proteins is our model system for developing a cell-free protein expression technology capable of producing virtually any membrane protein in a water-soluble functional form that is easy to study. We will synthesize ErbB-containing nanoparticles and characterize them for structure, function and for binding to peptide and small molecule libraries.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA155642-02
Application #
8298496
Study Section
Nanotechnology Study Section (NANO)
Program Officer
Fu, Yali
Project Start
2011-07-07
Project End
2016-05-31
Budget Start
2012-06-01
Budget End
2013-05-31
Support Year
2
Fiscal Year
2012
Total Cost
$318,513
Indirect Cost
$111,013
Name
University of California Davis
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618
Zhang, Hongyong; Li, Yuanpei; Lin, Tzu-Yin et al. (2014) Nanomicelle formulation modifies the pharmacokinetic profiles and cardiac toxicity of daunorubicin. Nanomedicine (Lond) 9:1807-20
He, Wei; Luo, Juntao; Bourguet, Feliza et al. (2013) Controlling the diameter, monodispersity, and solubility of ApoA1 nanolipoprotein particles using telodendrimer chemistry. Protein Sci 22:1078-86