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Nuclear magnetic resonance of the filamentous bacteriophage fd
Abstract:
The filamentous bacteriophage fd and its major coat protein are being studied by nuclear magnetic resonance (NMR) spectroscopy. 31P NMR shows that the chemical shielding tensor of the DNA phosphates of fd in solution is only slightly reduced in magnitude by motional averaging, indicating that DNA-protein interactions substantially immobilize the DNA packaged in the virus. There is no evidence of chemical interactions between the DNA backbone and the coat protein, since experiments on solid virus show the 31P resonances to have the same principle elements of its chemical shielding tensor as DNA. 1H and 13C NMR spectra of fd virus in solution indicate that the coat proteins are held rigidly in the structure except for some aliphatic side chains that undergo relatively rapid rotations. The presence of limited mobility in the viral coat proteins is substantiated by finding large quadrupole splittings in 2H NMR of deuterium labeled virions. The structure of the coat protein in a lipid environment differs significantly from that found for the assembled virus. Data from 1H and 13C NMR chemical shifts, amide proton exchange rates, and 13C relaxation measurements show that the coat protein in sodium dodecyl sulfate micelles has a native folded structure that varies from that of a typical globular protein or the coat protein in the virus by having a partially flexible backbone and some rapidly rotating aromatic rings.
Insights
Nuclear magnetic resonance (NMR) reveals filamentous bacteriophage fd DNA is immobilized by protein interactions. Coat proteins are rigid, but some side chains rotate, differing from their structure in lipid environments.
Area of Science:
- Structural biology
- Biophysics
- Molecular biology
Background:
- Filamentous bacteriophage fd is a model system for studying virus structure and dynamics.
- Nuclear magnetic resonance (NMR) spectroscopy is a powerful tool for investigating molecular structure and motion.
Purpose of the Study:
- To elucidate the structural dynamics of the fd virus and its coat protein using various NMR techniques.
- To compare the structure of the fd coat protein in the assembled virus with its structure in a lipid environment.
Main Methods:
- 31P, 1H, 13C, and 2H NMR spectroscopy were employed.
- Studies were conducted on fd in solution, as solid virus, and on coat protein in sodium dodecyl sulfate (SDS) micelles.
Main Results:
- DNA in fd virus is significantly immobilized by protein interactions, with no evidence of direct chemical interaction with the backbone.
- fd coat proteins in the virus are largely rigid, with some mobile side chains, confirmed by deuterium NMR.
- fd coat protein in SDS micelles exhibits a folded structure with a flexible backbone and rotating aromatic rings, differing from the viral structure.
Conclusions:
- DNA packaging within the fd virus leads to substantial immobilization.
- The fd coat protein maintains a native-like fold in micelles but exhibits distinct dynamic properties compared to the assembled virus.
- NMR spectroscopy provides detailed insights into the dynamic and structural differences of viral components in various environments.