title: Determination of 4-hydroxyproline-2-epimerase activity by capillary electrophoresis: A stereoselective platform for inhibitor screening of amino acid isomerases.
abstract: Isomerases involved in the metabolism of D/L-amino acids represent promising therapeutic targets for treatment of disease. Herein, we report a tunable platform for the assessment of enzymatic kinetics involving amino acid isomerization by CE that offers improved selectivity and sensitivity over traditional methods. Enzyme activity and competition assays were evaluated for various hydroxyproline diastereoisomers, proline enantiomers and their structural analogs using 4-hydroxyproline-2-epimerase as a model system. In this work, pyrrole 2-carboxylic acid was found to be a selective inhibitor of 4-hydroxyproline-2-epimerase with a half-maximal inhibition concentration of (2.3 + or - 0.1) mM. Reliable methods for unambiguous characterization of amino acid isomerases are required for the screening of novel inhibitors with epimerase and/or racemase activity.
authors.affiliation: Department of Chemistry and Chemical Biology, Hamilton, ON, Canada.
authors.forename: Jennilee M A
authors.initials: JM
authors.lastname: Gavina
authors.forename: Catharine E
authors.initials: CE
authors.lastname: White
authors.forename: Turlough M
authors.initials: TM
authors.lastname: Finan
authors.forename: Philip
authors.initials: P
authors.lastname: Britz-McKibbin
mesh_terms: D000591:Amino Acid Isomerases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D003550:Cystic Fibrosis / Q000473:pathology; D019075:Electrophoresis, Capillary / Q000379:methods; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006909:Hydroxyproline / Q000378:metabolism; D007700:Kinetics; D011392:Proline / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000472:pathogenicity; D012680:Sensitivity and Specificity
chemical_list: D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D011392:Proline; D000591:Amino Acid Isomerases; C021420:hydroxyproline epimerase; D006909:Hydroxyproline