The specificity of the interaction of bovine pancreatic ribonuclease A with natural and halogenated purine nucleotides.
The interaction between bovine pancreatic ribonuclease A (EC 3.1.4.22) and the purine nucleotides AMP, GMP, 6-chloropurine 5'-ribonucleotide and 8-bromoadenosine 5'-monophosphate was studied by u.v. difference spectroscopy. The stoicheiometry of the binding of the halogenated nucleotides to the enzyme shows a 1:1 ratio, as for the natural ones. The binding constants, Ka, for all four nucleotides at pH 5.5 were determined. They are within the same order of magnitude, though the nucleotides with a 6-amino group show a stronger interaction. The magnitude of the binding shows a reciprocal dependence on the ionic strength, which indicates an electrostatic interaction between ligand and enzyme. Finally, solvent-perturbation experiments show that all four nucleotides bind to the enzyme in a partially hydrophobic region. It is concluded that both halogenated and natural purine ribonucleotides interact in a similar manner with the enzyme molecule. The special synthesis and identification of 6-chloropurine 5'-ribonucleotide are discussed extensively. It is concluded that both halogenated and natural purine ribonucleotides interact in a similar manner with the enzyme molecule and thus the halogenated analogues are potential reagents for the affinity labelling of the purine-binding site.
D001665:Binding Sites; D055598:Chemical Phenomena; D002621:Chemistry; D011685:Purine Nucleotides / Q000138:chemical synthesis / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity
D011685:Purine Nucleotides; D012260:Ribonucleases