Probing the role of intercalating protein sidechains for kink formation in DNA

Abstract

Protein binding can induce DNA kinks, which are for example important to enhance the specificity of the interaction and to facilitate the assembly of multi protein complexes. The respective proteins frequently exhibit amino acid sidechains that intercalate between the DNA base steps at the site of the kink. However, on a molecular level there is only little information available about the role of individual sidechains for kink formation. To unravel structural principles of protein-induced DNA kinking we have performed molecular dynamics (MD) simulations of five complexes that varied in their architecture, function, and identity of intercalated residues. Simulations were performed for the DNA complexes of wildtype proteins (Sac7d, Sox-4, CcpA, TFAM, TBP) and for mutants, in which the intercalating residues were individually or combined replaced by alanine. The work revealed that for systems with multiple intercalated residues, not all of them are necessarily required for kink formation. In some complexes (Sox-4, TBP), one of the residues proved to be essential for kink formation, whereas the second residue has only a very small effect on the magnitude of the kink. In other systems (e.g. Sac7d) each of the intercalated residues proved to be individually capable of conferring a strong kink suggesting a partially redundant role of the intercalating residues. Mutation of the key residues responsible for kinking either resulted in stable complexes with reduced kink angles or caused conformational instability as evidenced by a shift of the kink to an adjacent base step. Thus, MD simulations can help to identify the role of individual inserted residues for kinking, which is not readily apparent from an inspection of the static structures. This information might be helpful for understanding protein-DNA interactions in more detail and for designing proteins with altered DNA binding properties in the future.

Atomic Resolution Insight into Sac7d Protein Binding to DNA and Associated Global Changes by Molecular Dynamics Simulations

“…[14] However,t he process of complex formation, possible intermediates,a nd how the large-scale DNAc onformational changes are triggered are not fully understood. [14] However,t he process of complex formation, possible intermediates,a nd how the large-scale DNAc onformational changes are triggered are not fully understood.…”

“…especially of the intercalating residues,f or the complex with DNAand acorrelation with the degree of induced bending. [14] However,t he process of complex formation, possible intermediates,a nd how the large-scale DNAc onformational changes are triggered are not fully understood.…”

Hydrophobic Amino Acids as Universal Elements of Protein-Induced DNA Structure Deformation

“…The intercalation of phenylalanine, leucine, or other hydrophobic residues into the DNA structure has been observed for several proteins that bind the DNA minor groove, including mitochondrial transcription factor A, male sex-determining factor SRY and bacterial catabolite control protein A. In these cases, the importance of the intercalated residues for modifying DNA structure was verified by molecular dynamics simulations of protein-DNA complexes in which these residues were replaced [18].…”

“…Another form of contacting the minor groove by a hydrophobic amino acid residue is intercalation of the residue into the DNA helix. During the binding of a TATA-box by the TBP, two phenylalanine residues intercalate into the DNA to stabilize the kink in the DNA double helix [16,18]. The intercalation of phenylalanine, leucine, or other hydrophobic residues into the DNA structure has been observed for several proteins that bind the DNA minor groove, including mitochondrial transcription factor A, male sex-determining factor SRY and bacterial catabolite control protein A.…”

Atomic Resolution Insight into Sac7d Protein Binding to DNA and Associated Global Changes by Molecular Dynamics Simulations

“…Simulations on the Sac7d–DNA also indicate that partial DNA bending can occur even without residue intercalation at the target site . Further in silico studies on Sac7d revealed the stabilizing role, especially of the intercalating residues, for the complex with DNA and a correlation with the degree of induced bending . However, the process of complex formation, possible intermediates, and how the large‐scale DNA conformational changes are triggered are not fully understood.…”