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Updated: Jun 23, 2026
Sedimentation Equilibrium of a Small Oligomer-forming Membrane Protein: Effect of Histidine Protonation on Pentameric Stability
Published on: April 2, 2015
Effect of coat protein mutations in bacteriophage fd studied by sedimentation analysis
A D Molina-Garcia1, S E Harding, F G Diaz
1University of Nottingham, Department of Biochemistry, University of Cambridge, CB2 1QW, United Kingdom.
Abstract:
(a) Bacteriophage fd is a filamentous virus that has previously been well characterized. (b) Earlier work using point mutagenesis indicated that a lysine residue at position 48 in the major coat protein plays a crucial role in interacting with the DNA and governing the assembly into an intact virion. (c) In this study the sedimentation properties (sedimentation velocity and equilibrium) of wild-type fd and two mutants substituted at lysine-48 (K48Q and K48A) were compared. (d) Both mutants are similar to each other [M(r) approximately (19.5 +/- 1.5) x 10(6)] but somewhat bigger than the wild-type [M(r) approximately (15.1 +/- 1.5) x 10(6)]. The value for the wild-type is consistent with earlier published values. (e) By combining these data with sedimentation coefficient data, it is possible to compare the contour lengths and relative flexibilities of the mutants with those of the wild-type virion. (f) The mutants are shown hydrodynamically to have larger contour lengths (as also observed by electron microscopy): the approximately 20% difference in values obtained assuming rigid particle hydrodynamics with those obtained from electron microscopy is strongly suggestive of some difference in flexibility between the wild-type and mutants.
Insights
Mutations in bacteriophage fd
Area of Science:
- Virology
- Molecular Biology
- Biophysics
Background:
- Bacteriophage fd is a well-characterized filamentous virus.
- Lysine-48 in the major coat protein is crucial for DNA interaction and virion assembly.
- Previous studies used point mutagenesis to identify this key residue.
Purpose of the Study:
- To compare the biophysical properties of wild-type fd with mutants at lysine-48.
- To investigate the role of lysine-48 in viral structure and assembly.
- To elucidate the hydrodynamic properties and flexibility of bacteriophage fd.
Main Methods:
- Sedimentation velocity and equilibrium analysis.
- Comparison of wild-type fd with K48Q and K48A mutants.
- Hydrodynamic modeling and electron microscopy data integration.
Main Results:
- Mutants K48Q and K48A exhibited similar molecular weights, larger than wild-type fd.
- Hydrodynamic analysis indicated increased contour lengths for the mutants.
- Electron microscopy corroborated the larger contour lengths observed in mutants.
Conclusions:
- The K48Q and K48A mutations alter the hydrodynamic properties of bacteriophage fd.
- Mutations at lysine-48 lead to increased viral contour length and potentially altered flexibility.
- These findings provide insights into the structural role of coat protein residues in filamentous phage assembly.
