DNA template-assisted inhibition of tyrosinase activity.
Enzyme-mediated polymerization reactions have been widely studied in the context of DNA template-assisted reactions. We have recently highlighted the ability of DNA templates to modulate enzyme-catalyzed single-step transformations. In this work, we focus on the intramolecular transformation of L-dopa catalyzed by enzyme tyrosinase and report a novel role of DNA templates in inhibition of the enzyme. The kinetics of mushroom tyrosinase monitored by UV-visible spectroscopy reveals significant decrease in the enzyme's efficiency in the presence of short double-stranded DNA molecules. KM of tyrosinase is found to increase by nearly 1.8-fold, implying a lower affinity of the enzyme for L-dopa, whereas Vmax is only marginally affected. The mode of inhibition is assessed to be a mixed mode with kinetic constants of inhibition in the micromolar range. Further, in the presence of cinnamic acid and DNA duplexes, the KM of tyrosinase increases nearly 3.5-fold, whereas Ki and Ki' are lowered by an order of magnitude. These results are a corollary of the known influence of substrate-template interactions and greater local substrate concentrations on enzyme activity and expand the paradigm with respect to use of DNA templates in enzyme-catalyzed reactions.
Department of Biological Engineering, Indian Institute of Technology Gandhinagar, VGEC Complex Chandkheda, Ahmedabad 382424, India.
Pallavi
P
Chilka
Department of Chemistry, Missouri State University, 901 S. National Avenue, Springfield, MO 65897, USA.
Sarah
S
Phillips
Department of Chemistry and Biological Engineering, Indian Institute of Technology Gandhinagar, VGEC Complex Chandkheda, Ahmedabad 382424, India. Electronic address: bdatta@iitgn.ac.in.
Bhaskar
B
Datta
D000363:Agaricales / Q000737:chemistry / Q000201:enzymology; D002934:Cinnamates / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D057075:Enzyme Assays; D005656:Fungal Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D007700:Kinetics; D007980:Levodopa / Q000737:chemistry*; D014442:Monophenol Monooxygenase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D009690:Nucleic Acid Conformation
D002934:Cinnamates; D005656:Fungal Proteins; C029010:cinnamic acid; D007980:Levodopa; D004247:DNA; D014442:Monophenol Monooxygenase
DNA-templated synthesis; Enzyme inhibition; Tyrosinase