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SPR analysis of promoter binding of Synechocystis PCC6803 transcription factors NtcA and CRP suggests cross‐talk and sheds light on regulation by effector molecules
Author(s) -
Forcada-Nadal Alicia,
Forchhammer Karl,
Rubio Vicente
Publication year - 2014
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2014.05.010
Subject(s) - promoter , transcription factor , biology , regulon , effector , dna binding protein , synechocystis , transcription (linguistics) , binding site , activator (genetics) , cyanobacteria , microbiology and biotechnology , gene , biochemistry , genetics , gene expression , bacteria , linguistics , philosophy
Surface plasmon resonance monitoring of the binding of transcription factors cAMP receptor protein (CRP) and nitrogen control factor of cyanobacteria (NtcA) from Synechocystis sp. PCC6803 to promoter fragments of glnA, glnN (NtcA regulon) and cccS (CRP regulon), revealed exclusive CRP binding to cccS, whereas NtcA was bound to all three promoters with different affinities, which were strongly increased by the NtcA activator 2‐oxoglutarate. Effective NtcA affinity for 2‐oxoglutarate varied with the promoter. High‐affinity promoters and the NtcA‐coactivating protein PII‐interacting protein X (PipX) increased NtcA affinity towards 2‐oxoglutarate, suggesting PipX‐stabilization of the 2‐oxoglutarate‐bound NtcA conformation. PipX binding to NtcA required 2‐oxoglutarate and was much tighter ( K d ≈ 85 nM) than to the PipX‐sequestering PII protein. NtcA appears to require more strongly PipX and 2‐oxoglutarate (2OG) for estimulating gene expression at promoters having “imperfect” NtcA binding sites.