Gene V Protein Dimerization and Cooperativity of Binding to Poly(dA)

Abstract

Gene V protein of bacteriophage f1 is a dimeric protein that binds cooperatively to single-stranded nucleic acids. In order to determine whether a monomer-dimer equilibrium has an appreciable effect upon the thermodynamics of gene V protein binding to nucleic acids, the dissociation constant for the protein dimer was investigated using size-exclusion chromatography. At concentrations ranging from 5 x 10(-10) to 1.2 x 10(-5) M, the Stokes radius of the protein was that expected of the dimer of the gene V protein. The Stokes radius of the protein was also independent of salt concentration from 0.2 to 1.0 M NaCl in a buffer containing 10 mM Tris-HCl, pH 7.4, and 1 mM EDTA. The binding of the dimeric gene V protein to poly(dA) was studied using a simplified lattice model for protein-protein interactions adapted for use with a dimeric protein that binds simultaneously to two strands of nucleic acid. Interpretation of the salt dependence, C = [d log(Kint omega)]/[d log(NaCl)], of binding of such a dimeric protein to nucleic acid using the theory of Record et al. (Record, M. T., et al. (1976) J. Mol. Biol. 107, 145-158) indicates that C is a function of the numbers of cations and anions released from protein and nucleic acid upon binding of the dimer, not of the monomer. Cooperativity of gene V protein binding to poly(dA) was studied with titration experiments that are sensitive to the degree of cooperativity of binding. The cooperativity factor omega, defined as the ratio of the binding constant for a site adjacent to a previously bound dimer to that for an isolated site, was found to be relatively insensitive to salt, with a value in the range of 2000-7000 for binding to poly(dA) at 3 degrees C and at 23 degrees C. This high cooperativity factor supports the suggestion that protein-protein contacts play a major role in the formation of the superhelical gene V protein-single-stranded nucleic acid complex.

Multimeric Self-assembly Equilibria Involving the Histone-like Protein H-NS

“…9A) (24,26). The interplay between oligomerization and DNA-binding properties of H-NS has been pointed out by several authors (14,22), and the control exerted by different ions on the H-NS oligomerization process could be seen as a general heterotropic effect on the DNA recognition process, as already described for other DNA-binding proteins (25,27).…”

Ultrafast Fluorescence Decay Profiles Reveal Differential Unstacking of 2-Aminopurine from Neighboring Bases in Single-Stranded DNA-Binding Protein Subsites

“…The labeled DNA was shifted to form only one major low-mobility band as g5p was added to give increasing N D /S molar ratios. The absence of 27 There was no evidence of the existence of more than one discrete type of complex. Saturation of the 44mers occurred at an N D /S ([g5p dimers]/[DNA strand]) molar ratio of ∼6 ( Figure 2B).…”

The 60-Residue C-Terminal Region of the Single-Stranded DNA Binding Protein of Herpes Simplex Virus Type 1 Is Required for Cooperative DNA Binding

“…The binding is typically cooperative, though the level of cooperativity varies widely. Much effort has been spent on elucidation of the structural mechanism accounting for the cooperativity and its functional implications in the case of the filamentous phage GVP (6,68), the phage T4 gp32 (10,33), the adenovirus DNA binding protein (DBP) (34), the Escherichia coli SSB (19,42), and the eukaryotic replication protein A (30,31), while little is known about the SSBs of the Herpesviridae.…”