Esterase

About: Esterase is a research topic. Over the lifetime, 7622 publications have been published within this topic receiving 168270 citations.

Membrane-permeable luciferin esters for assay of firefly luciferase in live intact cells.

Abstract: Five esters of luciferin were synthesized and compared with native luciferin as substrates for firefly luciferase expressed in live intact mammalian cells. The esters themselves were not substrates for purified luciferase, but four were substrates for a purified esterase and all appeared to be hydrolysed to luciferin within mammalian cells. At a substrate concentration of 0.01 mM, the peak luminescence from the cos cells expressing luciferase was up to 6-fold greater with the esters than with unmodified luciferin. At 0.1 mM, the difference between luciferin and the esters was decreased. The kinetics of the luminescent signal with the different luciferin esters varied significantly, indicating possible differences in the rates of uptake, breakdown and enzyme inhibition. The esters did not support luminescence from Escherichia coli cells expressing firefly luciferase, suggesting a lack of appropriate esterase activity in this particular strain. The esters could be useful for the assay of luciferase expression in intact mammalian cells when luciferin levels are limiting, for example in tissues, and in plants. Alternative luciferin derivatives may allow further improvements in sensitivity.

Biochemical analysis of a chlorfenapyr‐selected resistant strain of Tetranychus urticae Koch

Abstract: Tetranychus urticae Koch has recently developed resistance to chlorfenapyr in Australia and Japan, but no attempt has yet been made to describe the biochemical mechanisms involved in chlorfenapyr resistance. In this study a laboratory-selected chlorfenapyr-resistant strain was investigated. Resistance to chlorfenapyr was associated with a strong increase in esterase activity and P450 mono-oxygenase (MO) activity but a decrease in 3,3',5,5'-tetramethylbenzidine (TMBZ) peroxidation activity. Differences in esterase activities between susceptible and resistant strains increased with increasing carbon number of the aliphatic side-chain of the nitrophenol substrate. A 4.4-fold increase in the O-deethylation of 7-ethoxy-4-trifluoromethyl coumarin (7-EFC) mediated by P450 MOs was detected. Remarkably, the resistant strain showed only half of the total TMBZ peroxidation activity found in the susceptible strain. The activity of these enzymes was further determined on different crosses and back-crosses of both strains. Results indicated that activities correlated with chlorfenapyr susceptibility and could be considered as biochemical markers. Esterase isozymes of both strains and their crosses were separated with isoelectric focusing (IEF) and visualised after activity staining. It was clear that two distinct zones of enhanced esterase activity were present in the chlorfenapyr-resistant strain (EST 11, pI = 4.88 and EST 16, pI = 4.71). EST 11 was identified with inhibitors as a carboxylesterase. The relative presence and intensity of these esterase zones changed in the different crosses and could be seen as a marker for chlorfenapyr resistance. Glutathione-S-transferase and glucose-6-phosphate dehydrogenase activities were not significantly different between strains. A twofold decrease in TMBZ peroxidase activity in the resistant strain could reflect decreased activation of chlorfenapyr. On the basis of these results the involvement of P450 MOs and esterases in the activation and detoxification of chlorfenapyr in T. urticae is challenged and discussed.