Photo-cross-linking of CRP to nonspecific DNA in the absence of cAMP. DNA interacts with both the N- and C-terminal parts of the protein.
Adenosine cyclic 3',5'-phosphate receptor protein (CRP or CAP) is a regulatory protein involved in the transcription of several operons in Escherichia coli. cAMP-independent, nonspecific complexes of CRP and DNA were investigated by photochemical cross-linking of the protein to nonspecific DNA, whose thymines are substituted by 5-bromouracil (BrUra). The cross-linked protein was completely digested by trypsin, and the covalently bound peptides were sequenced. We identified two regions of the protein in close contact with DNA: one in the C-terminal part, overlapping the canonical helix-turn-helix motif, and the other one in the N-terminal part, which is usually not considered to belong to the DNA-interacting domain of CRP. This result lead us to propose models for nonspecific interaction, where the DNA is in contact with both the N- and C-terminal parts of the protein.
Centre de Biophysique Moléculaire, CNRS, Orléans, France.
M
M
Katouzian-Safadi
B
B
Blazy
J Y
JY
Cremet
J P
JP
Le Caer
J
J
Rossier
M
M
Charlier
D000595:Amino Acid Sequence; D001665:Binding Sites; D001976:Bromouracil / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D003432:Cross-Linking Reagents; D000242:Cyclic AMP / Q000494:pharmacology*; D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D008027:Light; D008969:Molecular Sequence Data; D010777:Photochemistry; D017433:Protein Structure, Secondary; D011953:Receptors, Cyclic AMP / Q000737:chemistry / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial; D011953:Receptors, Cyclic AMP; D001976:Bromouracil; D000242:Cyclic AMP; D014357:Trypsin