Ma. Sweezy et Sw. Morrical, SINGLE-STRANDED-DNA BINDING-PROPERTIES OF THE UVSY RECOMBINATION PROTEIN OF BACTERIOPHAGE-T4, Journal of Molecular Biology, 266(5), 1997, pp. 927-938
The uvsY protein is an essential component of the bacteriophage T4 gen
eral recombination machinery. The properties of this 16 kDa protein in
clude selective binding to ssDNA, as well as specific protein-protein
interactions with other T4 recombination proteins including uvsX (gene
ral recombinase) and gp32 (ssDNA-binding protein). uvsY promotes the a
ssembly of uvsX-ssDNA filaments, the active species in uvsX-catalyzed
DNA rearrangements, apparently by helping uvsX displace gp32 from the
ssDNA. To better understand the role of uvsY in the T4 recombination s
ystem, here we characterize the thermodynamic and molecular properties
of the interaction of the uvsY protein with a model single-stranded p
olynucleotide, epsilon DNA, which is a fluorescent, etheno-modified fo
rm of random-sequence ssDNA. We have found that the binding of uvsY pr
otein enhances the fluorescence of the epsilon DNA lattice and that th
e maximal amount of fluorescence enhancement observed is dependent on
salt concentration. In addition, we have used the epsilon DNA fluoresc
ence enhancement assay to establish thermodynamic parameters of bindin
g and to define some of the molecular details of uvsY-epsilon DNA inte
ractions. We show that uvsY binds to epsilon DNA in a non-cooperative
manner, with a binding site size of four nucleotide residues per monom
er of uvsY, and that this binding is salt-sensitive and involves the d
isplacement of anions from the uvsY protein. We further show that uvsY
protein binds preferentially to epsilon DNA over unmodified ssDNA. Th
e significance of these results is discussed in light of current model
s of uvsY action in the T4 recombination system. (C) 1997 Academic Pre
ss Limited.