Crystal structures of Y41H and Y41F mutants of gene V protein from Ff phage suggest possible protein-protein interactions in the GVP-ssDNA complex.
Gene V protein (GVP) encoded by the filamentous phage Ff (M13, fl, fd) is a homodimeric protein of 87 amino acids that binds to single-stranded DNA (ssDNA) nonspecifically and cooperatively. The structure (monoclinic C2 form) of the wild-type protein has been determined and refined at 1.8-A resolution [Skinner et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 2071-2075]. The monomer structure consists of a somewhat distorted five-stranded beta-barrel core with three prominent loops: a DNA-binding loop, a dyad loop, and a dimer contact loop. The amino acid residue at position 41 plays an important role in the dimer-dimer interactions of the protein-ssDNA complex. Two Y41 mutant structures have been studied by X-ray crystallography. The Y41F GVP structure has been refined to an R-factor of 0.180 at 2.2-A resolution and is very similar to the wild-type (wt) structure (rmsd of all C alpha atoms = 0.30 A). In contrast, Y41H GVP forms a new crystal lattice in the space group P2(1)2(1)2(1) with a = 77.18 A, b = 84.17 A, and c = 28.62 A. Its structure has been solved by the molecular replacement method and refined to an R-factor of 0.170 at 2.5-A resolution. The two monomers of Y41H are crystallographically independent, and their structures remain similar to wt-GVP but with significant differences, particularly in the DNA-binding hairpin region. In both crystals, the loop (residues 36-43) that contains the Y41 residue is involved in the crystal dimer packings but in a different manner. The dimer-dimer contacts found in the wt-GVP crystal may be important for GVP aggregation in the absence of DNA. In the presence of DNA, the dimer-dimer contacts may switch to the type found in the Y41H crystal, allowing the GVP-ssDNA complex to form cooperatively. A model of the complex, consistent with existing biochemical and biophysical data, has been constructed from those crystal packing data.
Guan
Y
Y
Biophysics Division, University of Illinois at Urbana-Champaign 61801.
Zhang
H
H
Konings
R N
RN
Hilbers
C W
CW
Terwilliger
T C
TC
Wang
A H
AH
D000595:Amino Acid Sequence; D017104:Bacteriophage M13; D018360:Crystallography, X-Ray; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry*; D005583:Fourier Analysis; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D014764:Viral Proteins / Q000737:chemistry*
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D014764:Viral Proteins; C069558:gene V protein, Enterobacteria phage f1