title: Recognition of ribonuclease A by 3'-5'-pyrophosphate-linked dinucleotide inhibitors: a molecular dynamics/continuum electrostatics analysis.
abstract: The proteins of the pancreatic ribonuclease A (RNase A) family catalyze the cleavage of the RNA polymer chain. The development of RNase inhibitors is of significant interest, as some of these compounds may have a therapeutic effect in pathological conditions associated with these proteins. The most potent low molecular weight inhibitor of RNase reported to date is the compound 5'-phospho-2'-deoxyuridine-3-pyrophosphate (P-->5)-adenosine-3-phosphate (pdUppA-3'-p). The 3',5'-pyrophosphate group of this compound increases its affinity and introduces structural features which seem to be unique in pyrophosphate-containing ligands bound to RNase A, such as the adoption of a syn conformation by the adenosine base at RNase subsite B(2) and the placement of the 5'-beta-phosphate of the adenylate (instead of the alpha-phosphate) at subsite P(1) where the phosphodiester bond cleavage occurs. In this work, we study by multi-ns molecular dynamics simulations the structural properties of RNase A complexes with the ligand pdUppA-3'-p and the related weaker inhibitor dUppA, which lacks the 3' and 5' terminal phosphate groups of pdUppA-3'-p. The simulations show that the adenylate 5'-beta-phosphate binding position and the adenosine syn orientation constitute robust structural features in both complexes, stabilized by persistent interactions with specific active-site residues of subsites P(1) and B(2). The simulation structures are used in conjunction with a continuum-electrostatics (Poisson-Boltzmann) model, to evaluate the relative binding affinity of the two complexes. The computed relative affinity of pdUppA-3'-p varies between -7.9 kcal/mol and -2.8 kcal/mol for a range of protein/ligand dielectric constants (epsilon(p)) 2-20, in good agreement with the experimental value (-3.6 kcal/mol); the agreement becomes exact with epsilon(p) = 8. The success of the continuum-electrostatics model suggests that the differences in affinity of the two ligands originate mainly from electrostatic interactions. A residue decomposition of the electrostatic free energies shows that the terminal phosphate groups of pdUppA-3'-p make increased interactions with residues Lys(7) and Lys(66) of the more remote sites P(2) and P(0), and His(119) of site P(1).
authors.affiliation: Department of Physics, University of Cyprus, Nicosia, Cyprus.
authors.forename: Savvas
authors.initials: S
authors.lastname: Polydoridis
authors.forename: Demetres D
authors.initials: DD
authors.lastname: Leonidas
authors.forename: Nikos G
authors.initials: NG
authors.lastname: Oikonomakos
authors.forename: Georgios
authors.initials: G
authors.lastname: Archontis
mesh_terms: D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000737:chemistry; D001665:Binding Sites; D003198:Computer Simulation; D003856:Deoxyuracil Nucleotides / Q000737:chemistry*; D008024:Ligands; D011487:Protein Conformation; D012259:Ribonuclease, Pancreatic / Q000037:antagonists & inhibitors / Q000737:chemistry*
chemical_list: C528963:5'-phospho-2'-deoxyuridine-3-pyrophosphate (P-5)-adenosine-3-phosphate; D003856:Deoxyuracil Nucleotides; D008024:Ligands; D000249:Adenosine Monophosphate; D012259:Ribonuclease, Pancreatic