X-ray diffraction analysis of gene V protein encoded by filamentous bacteriophage Ff

Abstract

The gene V protein encoded by bacteriophage Ff is a single-stranded DNA binding protein and it plays an important role in the replication cycle of the phage. The crystal structure of the gene V protein was determined using multiwavelength anomalous diffraction on the selenomethionine-containing wild-type and isoleucine-47 $\to$ methionine mutant proteins with x-ray diffraction data phased to 2.5 A resolution. The structure of the wild-type protein was refined to an R factor of 19.1% using native data to 1.8 A resolution. The gene V protein monomer is largely composed of $\beta$-structures, including a distorted five stranded antiparallel $\beta$-barrel and two prominent extended $\beta$-hairpins. The two monomers are closely associated together to form a dimer. The DNA binding site of the protein was explored by the qualitative electrostatic potential calculations. The result from the preliminary x-ray diffraction analysis of co-crystals of gene V protein and oligonucleotides was also presented.

The crystal structures of four mutant gene V proteins were solved and refined to $\sim$2 A resolutions. Three of these mutants were apolar substitutions in the hydrophobic core of the protein. One mutant, Arg 82 $\to$ Cys, is at the protein surface and is involved in breaking up of a surface salt bridge. The effects of these substitutions on the stability of the protein were discussed.