title: Sensitization of the Escherichia coli cyclic AMP receptor protein to trypsin cleavage by polydeoxyribonucleotides and polyribonucleotides.
abstract: In the absence of cAMP the cyclic AMP receptor protein (CRP) is relatively resistant to trypsin whereas the cAMP X CRP complex is attacked yielding N-terminal core fragments of 14,300 and 18,500 Da which still bind cAMP. The DNA X CRP complex formed at low ionic strength in the absence of cAMP is cleaved by trypsin with the formation of 9,700- and 6,000-Da fragments and the concomitant loss of cAMP binding activity. DNA X CRP remains as resistant to attack by subtilisin, clostripain, and the Staphylococcus aureus V8 protease as unliganded CRP but is slowly digested by chymotrypsin. All of the double-stranded polydeoxyribonucleotides and several of the single-stranded polydeoxyribonucleotides and polyribonucleotides tested render CRP sensitive to cleavage by trypsin. CRP is less rapidly cleaved by trypsin in the presence of d(A)n, d(I)n, and r(C)n indicative of a weaker affinity of CRP for these polynucleotides. The 9,700-Da fragment is N-terminal in CRP and probably terminates at Lys-89. The loss of cAMP binding activity following trypsin cleavage of DNA X CRP indicates that regions beyond this residue are important in the function of the cAMP-binding domain of CRP. The 6,000-Da fragment extends from Val-131 to Arg-185 or Lys-188 and contains part of the F helix involved in DNA binding by CRP.
mesh_terms: D000595:Amino Acid Sequence; D000242:Cyclic AMP / Q000378:metabolism; D004926:Escherichia coli / Q000032:analysis*; D009994:Osmolar Concentration; D010446:Peptide Fragments / Q000032:analysis; D011089:Polydeoxyribonucleotides / Q000494:pharmacology*; D011131:Polyribonucleotides / Q000494:pharmacology*; D011953:Receptors, Cyclic AMP / Q000378:metabolism*; D013997:Time Factors; D014357:Trypsin / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D011089:Polydeoxyribonucleotides; D011131:Polyribonucleotides; D011953:Receptors, Cyclic AMP; D000242:Cyclic AMP; D014357:Trypsin