SINGLE-STRANDED-DNA BINDING-PROTEIN FROM BACTERIOPHAGE-CF - CHARACTERIZATION, GENE LOCALIZATION AND PROTEIN-SSDNA COMPLEX

The single-stranded DNA binding protein from the filamentous bacteriop hage cf has been purified and characterized. The first 12 amino acids, resulting from the N-terminal amino acid sequencing analysis of the p rotein, agree with an open reading frame (ORF) on the cf genome. The O RF contains 294 bp and codes for a 98 a.a. protein of molecular weight 10.8 kDa, consistent with the result from the denaturing protein gel analysis. The protein appears to be a homodimer as evident from the ap parent molecular weight of about 22 kDa obtained from native protein g el analysis. The gene location of the protein has been identified as g ene V of the cf single stranded genome, same as that from the M13 phag e. The GVP of cf shows a strong sequence homology to the ssDNA binding proteins of Ff, IKe and Pf3 filamentous phages. The DNA binding wing of GVP, conserved among the filamentous phages, has been predicted for cf, To further characterize the protein, the GVP-ssDNA complex of cf has been purified from the infected host (Xanthomonas campestris pv. c itri) by density gradient centrifugation, Transmission electron micros copy (TEM) images of the complex showed that it is about 1200 nm in le ngth and 9 nm in diameter and it has a highly regular morphology with a central groove shadow running along the entire structure, but withou t any apparent helical pattern seen in the M13 complex, The GVP-ssDNA complex of cf seems more rigid than that of M13. Our computer modeling study suggested that this difference between the two complexes may be due to the additional 11 or 12 amino acids at the C-terminal end of t he cf-GVP.