Protein p6 from Bacillus subtilis phage phi29 binds double-stranded DNA, forming a large nucleoprotein complex all along the viral genome, and has been proposed to be an architectural protein with a global role in genome organization. Here, we have characterized quantitatively the DNA binding properties of protein p6 by means of sedimentation velocity and sedimentation equilibrium experiments permitting determination of the strength and stoichiometry of complex formation. The composition dependence of protein binding to DNA is quantitatively consistent with a model in which the protein undergoes a reversible monomer-dimer self-association, and the dimeric species binds noncooperatively to the DNA. We also have found that when the anisotropic bendability periodicity of the nucleotide sequence preferred by p6 is modified, nucleocomplex formation is impaired. In addition, suppression of complex formation at high ionic strength is reversed by the addition of high concentrations of an inert polymer, mimicking the crowded intracellular environment. The results obtained in this work illustrate how macromolecular crowding could act as a metabolic buffer that can significantly extend the range of intracellular conditions under which a specific reaction may occur.