Role of DNA conformation & energetic insights in Msx-1-DNA recognition as revealed by molecular dynamics studies on specific and nonspecific complexes

<div><p>In most of homeodomain–DNA complexes, glutamine or lysine is present at 50th position and interacts with 5th and 6th nucleotide of core recognition region. Molecular dynamics simulations of Msx-1–DNA complex (Q50-TG) and its variant complexes, that is specific (Q50K-CC), nonspecific (Q50-CC) having mutation in DNA and (Q50K-TG) in protein, have been carried out. Analysis of protein–DNA interactions and structure of DNA in specific and nonspecific complexes show that amino acid residues use sequence-dependent shape of DNA to interact. The binding free energies of all four complexes were analysed to define role of amino acid residue at 50th position in terms of binding strength considering the variation in DNA on stability of protein–DNA complexes. The order of stability of protein–DNA complexes shows that specific complexes are more stable than nonspecific ones. Decomposition analysis shows that N-terminal amino acid residues have been found to contribute maximally in binding free energy of protein–DNA complexes. Among specific protein–DNA complexes, K50 contributes more as compared to Q50 towards binding free energy in respective complexes. The sequence dependence of local conformation of DNA enables Q50/Q50K to make hydrogen bond with nucleotide(s) of DNA. The changes in amino acid sequence of protein are accommodated and stabilized around TAAT core region of DNA having variation in nucleotides.</p></div

Molecular dynamics simulations show altered secondary structure of clawless in binary complex with DNA providing insights into aristaless-clawless-DNA ternary complex formation

<p>Aristaless (Al) and clawless (Cll) homeodomains that are involved in leg development in <i>Drosophila melanogaster</i> are known to bind cooperatively to 5′-(T/C)TAATTAA(T/A)(T/A)G-3′ DNA sequence, but the mechanism of their binding to DNA is unknown. Molecular dynamics (MD) studies have been carried out on binary, ternary, and reconstructed protein–DNA complexes involving Al, Cll, and DNA along with binding free energy analysis of these complexes. Analysis of MD trajectories of Cll–3A01, binary complex reveals that C-terminal end of helixIII of Cll, unwind in the absence of Al and remains so in reconstructed ternary complex, Cll–3A01–Al. In addition, this change in secondary structure of Cll does not allow it to form protein–protein interactions with Al in the ternary reconstructed complex. However, secondary structure of Cll and its interactions are maintained in other reconstructed ternary complex, Al–3A01–Cll where Cll binds to Al–3A01, binary complex to form ternary complex. These interactions as observed during MD simulations compare well with those observed in ternary crystal structure. Thus, this study highlights the role of helixIII of Cll and protein–protein interactions while proposing likely mechanism of recognition in ternary complex, Al–Cll–DNA.</p

Hydrogen bonding between ligand and protein active site residues.

<p>Dash line shows hydrogen bond between, trimethoprim (magenta) and protein active site residues (green), A) TMP-sensitive-Dfr-TMP complex, B) PGB01-DfrA7-TMP complex.</p

Effect of microwave radiation on cell viability.

<p>A. Thermal effect of microwave radiation: Temperature of LB vs. time of exposure; B. Viability of cells vs. time of microwave exposure.</p

Diagram of amplified class 1 integron.

<p>Schematic representation of class 1 integron amplicon of the strain PGB01 amplified with the Int₂F-3′CS primer pair. Numbers correspond to sequence positions in EMBL Ac. No. FN563072.</p

Superimposition of protein active site residues.

<p>Superimposed side chain representation of protein active site residues of TMP-sensitive-Dfr (magenta) and PGB01-DfrA7 (orange).</p