Mode of DNA binding with γ-butyrolactone receptor protein CprB from Streptomyces coelicolor revealed by site-specific fluorescence dynamics.
BACKGROUND
The γ-butyrolactone (GBL) binding transcription factors in Streptomyces species are known for their involvement in quorum sensing where they control the expression of various genes initiating secondary metabolic pathways. The structurally characterized member of this family CprB from Streptomyces coelicolor had earlier been demonstrated to bind a multitude of sequences containing a specific binding signature. Though structural breakthrough has been obtained for its complex with a consensus DNA sequence there is, however a dearth of information regarding the overall and site specific dynamics of protein-DNA interaction.


METHODS
To delineate the effect of CprB on the bound DNA, changes in motional dynamics of the fluorescent probe 2-aminopurine were monitored at three conserved base positions (5th, 12th and 23rd) for two DNA sequences: the consensus and the biologically relevant cognate element, on complex formation.


RESULTS
The changes in lifetime and generalized order parameter revealed a similarity in the binding pattern of the protein to both sequences with greater dynamic restriction at the end positions, 5th and 23rd, as compared to the middle 12th position. Also differences within this pattern demonstrated the influence of even small changes in sequence on protein interactions.


CONCLUSIONS
Here the study of motional dynamics was instrumental in establishing a structural footprint for the cognate DNA sequence and explaining the dynamics for the consensus DNA from structural correspondence.


CONCLUSIONS
Motional dynamics can be a powerful tool to efficiently study the mode of DNA binding to proteins that interact differentially with a plethora of DNA sequences, even in the absence of structural breakthrough.
Department of Chemistry, Indian Institute of Technology Bombay, Mumbai, Maharashtra 400076, India.
Anwesha
A
Biswas
Department of Chemical Sciences, Tata Institute of Fundamental Research, Mumbai, Maharashtra 400005, India.
Satya
S
Narayan
Department of Chemical Sciences, Tata Institute of Fundamental Research, Mumbai, Maharashtra 400005, India.
Mamata V
MV
Kallianpur
Department of Chemical Sciences, Tata Institute of Fundamental Research, Mumbai, Maharashtra 400005, India. Electronic address: gk@tifr.res.in.
G
G
Krishnamoorthy
Department of Chemistry, Indian Institute of Technology Bombay, Mumbai, Maharashtra 400076, India. Electronic address: ruchi@chem.iitb.ac.in.
Ruchi
R
Anand
D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D005453:Fluorescence; D008969:Molecular Sequence Data; D011963:Receptors, GABA-A / Q000378:metabolism*; D048372:Streptomyces coelicolor / Q000378:metabolism*
C037481:4-butyrolactone receptor; D011963:Receptors, GABA-A; D004247:DNA
2-Aminopurine; Anisotropy; Fluorescence lifetime; γ-Butyrolactone