Structure and Flexibility Adaptation in Nonspecific and Specific Protein-DNA Complexes

Abstract

Interaction of regulatory DNA binding proteins with their target sites is usually preceded by binding to nonspecific DNA. This speeds up the search for the target site by several orders of magnitude. We report the solution structure and dynamics of the complex of a dimeric lac repressor DNA binding domain with nonspecific DNA. The same set of residues can switch roles from a purely electrostatic interaction with the DNA backbone in the nonspecific complex to a highly specific binding mode with the base pairs of the cognate operator sequence. The protein-DNA interface of the nonspecific complex is flexible on biologically relevant time scales that may assist in the rapid and efficient finding of the target site.

Extrinsic Interactions Dominate Helical Propensity in Coupled Binding and Folding of the Lactose Repressor Protein Hinge Helix

“…For LacI and PurR, the hinge folds into a helix only upon cognate operator binding, whereas this region remains unstructured in other states, apparently even when bound to nonspecific DNA (14,15,(19)(20)(21)23,24,66,67). The hinge helix participates in multiple interfaces ( Figure 1B) -between the partner hinge helices within the dimer, between the hinge helix and the top surfaces of the core N-subdomains, and between the minor groove of DNA and side chains in the hinge helix (25,68).…”

“…In NMR structures of the truncated protein alone or with nonspecific DNA the hinge is disordered, but folded in the presence of operator DNA (19-21). Thus, in addition to its importance in highaffinity DNA binding and inducer response, the hinge region may also be part of a switch between nonspecific and specific operator binding (14,15,(19)(20)(21)(22).Residues of the hinge helix participate both in high affinity operator-binding and in allosteric communication between the core inducer binding site and the DNA binding domain (14,15,21,23,24). This helix interacts with minor-groove bases through side chains of Asn50, Ala53, Gln54, Leu56, and Ala57 ( Figure 1B) (25).…”

Protein–DNA Electrostatics

“…Non-specific interactions between proteins and DNA are poorly documented, but the predominance of electrostatic undeniable [14,15,16,17,18,19]. Proteins that bind to DNA are most often positively charged.…”

“…However, it is tempting to ask whether this particular protein shape may play a role in non specific protein-DNA interactions, at work during the target sequence search. In this regard, it is interesting to note that structural studies of some non-specific protein-DNA complexes show a gap between the two macromolecules, filled with solvent [14,15,17,18,19]. This observation suggests the existence of a force that counteracts the electrostatic attraction.…”

“…This supports the idea that electrostatics is involved to some extent in the intermittent behavior, with a probable role for the dissolved salt ions. Electrostatics plays indeed a major role in the protein-DNA interaction [14,15,16,17,18,19,20]. The reason is that DNA is very strongly negatively charged (-2 charges per base pair).…”