In this section
@article{McFarlane:2020:10.21769/bioprotoc.3599,
author = {McFarlane, C and Murray, J},
doi = {10.21769/bioprotoc.3599},
journal = {Bio-protocol},
pages = {1--10},
title = {A sensitive coupled enzyme assay for measuring kinase and ATPase kinetics using ADP-specific hexokinase},
url = {http://dx.doi.org/10.21769/bioprotoc.3599},
volume = {10},
year = {2020}
}
TY - JOUR
AB - Kinases and ATPases perform essential biological functions in metabolism and regulation.Activity of these enzymes is commonly measured by coupling ATP consumption to the synthesis of adetectable product. For most assay systems the ATP concentration during the reaction is unknown,compromising the precision of the assay. Using the ADP-specific hexokinase (ADP-HK) from the thermophilic archaeon Thermococcus litoralisthe protocol outlined here allows real time coupling of ATP consumption to downstream signal changeenabling accurate kinetic measurements. ADP-HK phosphorylates glucose that is then used by glucose6-phosphate dehydrogenase to reduce NAD+ to NADH which can be measured at 340 nm. We haveshown this assay to be sensitive to the detection of micromole quantities of ADP with no detectablebackground from ATP.
AU - McFarlane,C
AU - Murray,J
DO - 10.21769/bioprotoc.3599
EP - 10
PY - 2020///
SN - 2331-8325
SP - 1
TI - A sensitive coupled enzyme assay for measuring kinase and ATPase kinetics using ADP-specific hexokinase
T2 - Bio-protocol
UR - http://dx.doi.org/10.21769/bioprotoc.3599
UR - https://bio-protocol.org/e3599
VL - 10
ER -
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