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39738-B4
Studies to Investigate the Protein-Based Molecular Recognition of the Adenine Ring
Year 4 of this grant was a result of a time extension from the ACS-PRF. The PI is grateful for this additional year as it allowed him to involve three additional students in research and make additional progress toward completing the work outlined in the proposal. Most of the work in Year 4 was the design and synthesis of novel aromatic compounds based on the pyrimidine ring. These compounds were designed to take advantage of binding opportunities in the adenine-binding region of the model enzyme, APH(3')-IIIa (AIM #3). Three undergraduate research students were involved in the synthesis and testing of this series of molecules. The pyrimidine-based compounds were synthesized from various substituted acetic acids via a vinamidinium salt . The substituent (R) on the acetic acetic acid provides the 5-position substituent after elaboration to the pyrimidine with guanidine-hydrochloride. The phenyl substituents include bromo, methyl and methoxy groups in the para position. The 2-amino-5-chloropyrimidine is available commercially, but was synthesized (less costly) for comparison. Yields have been low (13-36%), but sufficient quantities for characterization and evaluation were easily obtained.
An electrostatic potential surface was calculated (at the B3LYP/6-31G** level of theory) for each of these compounds and tested as an inhibitor of APH(3')-IIIa. Taken together, the inhibitory data, along with the electrostatic potential surfaces, provided new details on the molecular and electrostatic determinants for small, aromatic systems to be inhibitors of this model kinase. Briefly, a compound of this type has to have the necessary combination of H-bond acceptors/donors or have some combination of these acceptors/donors along with an electron deficient aromatic system that may promote the pi-pi stacking interaction that is known to form in the adenine-binding region of APH(3')-IIIa. The results indicate that the inhibitory constants decreased (more potent inhibitors) as the pyrimidine ring of these compounds became more electron deficient. The quantum-mechanical calculations also revealed that the partial charges on N atoms, and H atoms bonded to N atoms, in this series of molecules did not change significantly. Therefore, the H-bonding potential of these compounds were not dramatically altered and that inhibitory efficacy can be related to the ability to form the pi-pi stacking interaction in the active site of the enzyme. Although this grant has concluded, this work will continue temporarily with internal funds until more external funding can be obtained.
