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Three-dimensional structure of a cloning vector. X-ray diffraction studies of filamentous bacteriophage M13 at 7 A
M J Glucksman1, S Bhattacharjee, L Makowski
1Department of Biochemistry and Molecular Biophysics, College of Physicians and Surgeons of Columbia University, New York, NY 10032.
Abstract:
Filamentous bacteriophage M13 is a single-stranded DNA phage about 65 A in diameter and 9300 A long. X-ray diffraction studies of magnetically oriented fibers of native, mercury and iodine-labeled phage particles have been used to determine the arrangement of the major coat protein, the gene 8 product, in the virion. The coat protein is made up of a single gently curving alpha-helix extending from approximately Pro6 to near the carboxyl terminus. The axis of the alpha-helix is tilted about 20 degrees from the viral axis and wraps around the axis in a right-handed helical sense. The surface of the virus is made up largely of polar residues in the amino-terminal half of the protein including the segment of alpha-helix extending from Pro6 to Tyr24. The interior surface of the protein coat faces the DNA and consists of an amphipathic helical segment extending from Thr36 to Ser50. The alpha-helices form a tightly packed 15 to 20 A thick cylindrical coat around the DNA. This structural model provides insight into the potential sites for incorporating foreign protein domains that may act as functional binding sites on the surface of M13.
Insights
Filamentous bacteriophage M13
Area of Science:
- Structural biology
- Virology
- Biophysics
Background:
- Filamentous bacteriophage M13 is a single-stranded DNA virus.
- The major coat protein (gene 8 product) forms the virion structure.
- Understanding M13 structure is key for phage display applications.
Purpose of the Study:
- Determine the arrangement of the M13 major coat protein.
- Elucidate the helical structure and orientation within the virion.
- Identify potential sites for foreign protein insertion.
Main Methods:
- X-ray diffraction studies of magnetically oriented phage fibers.
- Analysis of native, mercury, and iodine-labeled M13 particles.
- High-resolution structural modeling of the coat protein.
Main Results:
- The M13 coat protein forms a gently curving alpha-helix.
- This alpha-helix is tilted ~20 degrees from the viral axis.
- The protein coat is a tightly packed cylinder (~15-20 A thick) around the DNA.
Conclusions:
- A detailed structural model of the M13 virion coat protein is established.
- The model highlights polar residues on the outer surface and amphipathic helices facing the DNA.
- The findings offer insights into engineering M13 for novel binding functionalities.