Title | Conservative Secondary Shell Substitution In Cyclooxygenase-2 Reduces Inhibition by Indomethacin Amides and Esters via Altered Enzyme Dynamics. |
Publication Type | Journal Article |
Year of Publication | 2016 |
Authors | Konkle ME, Blobaum AL, Moth CW, Prusakiewicz JJ, Xu S, Ghebreselasie K, Akingbade D, Jacobs AT, Rouzer CA, Lybrand TP, Marnett LJ |
Journal | Biochemistry |
Volume | 55 |
Issue | 2 |
Pagination | 348-59 |
Date Published | 2016 Jan 19 |
ISSN | 1520-4995 |
Keywords | Amides, Computational Biology, Cyclooxygenase 2, Enzyme Activation, Esters, Indomethacin, Protein Structure, Secondary, Structure-Activity Relationship |
Abstract | The cyclooxygenase enzymes (COX-1 and COX-2) are the therapeutic targets of nonsteroidal anti-inflammatory drugs (NSAIDs). Neutralization of the carboxylic acid moiety of the NSAID indomethacin to an ester or amide functionality confers COX-2 selectivity, but the molecular basis for this selectivity has not been completely revealed through mutagenesis studies and/or X-ray crystallographic attempts. We expressed and assayed a number of divergent secondary shell COX-2 active site mutants and found that a COX-2 to COX-1 change at position 472 (Leu in COX-2, Met in COX-1) reduced the potency of enzyme inhibition by a series of COX-2-selective indomethacin amides and esters. In contrast, the potencies of indomethacin, arylacetic acid, propionic acid, and COX-2-selective diarylheterocycle inhibitors were either unaffected or only mildly affected by this mutation. Molecular dynamics simulations revealed identical equilibrium enzyme structures around residue 472; however, calculations indicated that the L472M mutation impacted local low-frequency dynamical COX constriction site motions by stabilizing the active site entrance and slowing constriction site dynamics. Kinetic analysis of inhibitor binding is consistent with the computational findings. |
DOI | 10.1021/acs.biochem.5b01222 |
Alternate Journal | Biochemistry |
PubMed ID | 26704937 |
PubMed Central ID | PMC4721528 |
Grant List | R01 CA089450 / CA / NCI NIH HHS / United States CA89450 / CA / NCI NIH HHS / United States T32GM65086-02 / GM / NIGMS NIH HHS / United States |
Submitted by kej2006 on June 6, 2018 - 4:11pm