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Apolipoprotein C-III Nanodiscs Studied by Site-Specific Tryptophan Fluorescence
Chase A Brisbois1, Jennifer C Lee1
1Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health , Bethesda, Maryland 20892, United States.
Biochemistry
|August 17, 2016
Summary
Apolipoprotein C-III (ApoC-III) nanodisc formation is influenced by tryptophan residue W42, crucial for lipid binding and assembly. This finding highlights W42
Area of Science:
- Biochemistry
- Structural Biology
- Lipid Metabolism
Background:
- Apolipoprotein C-III (ApoC-III) is a key component of high-density lipoproteins (HDLs).
- ApoC-III facilitates the remodeling of lipid vesicles into HDL-like particles called nanodiscs.
- Understanding ApoC-III's structure-function relationship in nanodiscs is vital for lipid metabolism research.
Purpose of the Study:
- To investigate the local side chain environments and interactions of ApoC-III within nanodiscs.
- To characterize the role of specific tryptophan residues (W42, W54, W65) in nanodisc formation and stability.
- To compare ApoC-III's binding environment in nanodiscs to micellar and vesicle states.
Main Methods:
- Utilized single-tryptophan (Trp) containing ApoC-III mutants.
- Employed transmission electron microscopy (TEM) for ultrastructural characterization.
- Applied circular dichroism (CD) spectroscopy for secondary structure analysis.
- Performed time-resolved anisotropy measurements to assess Trp side chain dynamics.
Main Results:
- Nanodiscs formed were structurally consistent (15 ± 2 nm diameter, ~25 proteins/particle, 45-51% helicity).
- W54 exhibited the most hydrophobic environment, followed by W42 and W65.
- Trp side chain mobility was uncorrelated with environmental polarity; W54 was most mobile.
- ApoC-III binding in nanodiscs resembles a bilayer-bound state.
- W42 demonstrated the most wild-type-like behavior during nanodisc formation across temperatures.
Conclusions:
- The interfacial location of W42 is critical for lipid binding and nanodisc assembly.
- W42's invariance among mammals suggests its biological significance in ApoC-III function.
- Modest mutations at Trp sites can reveal important insights into protein-lipid interactions.
