A pyrophosphatase-coupled assay to monitor the NTase activity of cGAS

2019 ◽  
pp. 77-86 ◽  
Author(s):  
Richard Hooy ◽  
Jungsan Sohn
Keyword(s):  
Coupled Assay

scholarly journals An enzyme-coupled assay for acyl-CoA synthetase.

1991 ◽  
Vol 32 (10) ◽  
pp. 1709-1712 ◽  
Author(s):  
K Ichihara ◽  
Y Shibasaki
Keyword(s):  
Coupled Assay

An enzyme-coupled assay enables rapid protein engineering for geraniol production in yeast

2018 ◽  
Vol 139 ◽  
pp. 95-100 ◽  
Author(s):  
Jyun-Liang Lin ◽  
Holly Ekas ◽  
Kelly Markham ◽  
Hal S. Alper
Keyword(s):  
Protein Engineering ◽  
Coupled Assay

scholarly journals Establishment of live-cell-based coupled assay system for identification of compounds to modulate metabolic activities of cells

Cell Reports ◽  
2021 ◽  
Vol 36 (1) ◽  
pp. 109311
Author(s):  
Kouichi Yanagi ◽  
Toru Komatsu ◽  
Shusuke Ogihara ◽  
Takayoshi Okabe ◽  
Hirotatsu Kojima ◽  
...  
Keyword(s):  
Live Cell ◽  
Assay System ◽  
Metabolic Activities ◽  
Coupled Assay

scholarly journals A Direct Pyrophosphatase-coupled Assay Provides New Insights into the Activation of the Secreted Adenylate Cyclase fromBordetella pertussisby Calmodulin

2002 ◽  
Vol 277 (25) ◽  
pp. 22289-22296 ◽  
Author(s):  
Anthony J. Lawrence ◽  
John G. Coote ◽  
Yasmin F. Kazi ◽  
Paul D. Lawrence ◽  
Julia MacDonald-Fyall ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Coupled Assay

Coupled assay for uroporphyrinogen III synthase

1997 ◽  
pp. 327-336 ◽  
Author(s):  
Peter M. Shoolingin-Jordan ◽  
Robert Leadbeater
Keyword(s):  
Coupled Assay

Determination of tissue plasminogen activator and its "fast" inhibitor in plasma.

1986 ◽  
Vol 32 (3) ◽  
pp. 482-485 ◽  
Author(s):  
J Chmielewska ◽  
B Wiman
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Plasma Samples ◽  
Inhibitor Concentration ◽  
Blood Samples ◽  
Analytical Recovery ◽  
Specific Determination ◽  
Tissue Plasminogen ◽  
Coupled Assay

Abstract We describe efficient, accurate methods for specific determination of tissue plasminogen activator (t-PA, EC 3.4.21.31) and its "fast" inhibitor in plasma. In this coupled assay, a sample containing t-PA is incubated with plasminogen, a plasmin (EC 3.4.21.7) substrate of low Km and high Kcat, and fibrin as a stimulator. The inhibitor of t-PA is determined by incubating the sample with a known amount of t-PA in excess, then determining the residual t-PA. Both t-PA and t-PA inhibitor can be determined in many samples simultaneously within a few hours. These assays are modifications of procedures described by us (Clin Chim Acta 1983;127:279-88 and Thromb Res 1983;31:427-36). Their accuracy as assessed by analytical recovery of pure t-PA added to blood samples (91 +/- 4%) or of partly purified inhibitor added to plasma samples (102 +/- 10%) is satisfactory, as is their precision. For the t-PA assay the CV was 1.6% (within run) or 4.1% (between run). The corresponding values for the inhibitor assay were 4.5% (within run) or 8.4% (between run) if the inhibitor concentration exceeded 3 arb. units/mL.

Kinetic and mechanistic analyses of new classes of inhibitors of two-component signal transduction systems using a coupled assay containing HpkA–DrrA from Thermotoga maritima

Microbiology ◽  
2004 ◽  
Vol 150 (4) ◽  
pp. 885-896 ◽  
Author(s):  
J. Estelle Foster ◽  
Qin Sheng ◽  
Jonathan R. McClain ◽  
Mark Bures ◽  
Thalia I. Nicas ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Histidine Kinase ◽  
Competitive Inhibition ◽  
Thermotoga Maritima ◽  
Phosphorylation Site ◽  
Phosphoryl Group ◽  
Two Component ◽  
Two Component Signal Transduction ◽  
Coupled Assay ◽  
Signal Transduction Systems

Two-component signal transduction systems (TCSs) play fundamental roles in bacterial survival and pathogenesis and have been proposed as targets for the development of novel classes of antibiotics. A new coupled assay was developed and applied to analyse the kinetic mechanisms of three new kinds of inhibitors of TCS function. The assay exploits the biochemical properties of the cognate HpkA–DrrA histidine kinase–response regulator pair from Thermotoga maritima and allows multiple turnovers of HpkA, linear formation of phosphorylated DrrA, and Michaelis–Menten analysis of inhibitors. The assay was validated in several ways, including confirmation of competitive inhibition by adenosine 5′-β,γ-imidotriphosphate (AMP-PNP). The coupled assay, autophosphorylation and chemical cross-linking were used to determine the mechanisms by which several compounds inhibit TCS function. A cyanoacetoacetamide showed non-competitive inhibition with respect to ATP concentration in the coupled assay. The cyanoacetoacetamide also inhibited autophosphorylation of histidine kinases from other bacteria, indicating that the coupled assay could detect general inhibitors of histidine kinase function. Inhibition of HpkA autophosphorylation by this compound was probably caused by aggregation of HpkA, consistent with a previous model for other hydrophobic compounds. In contrast, ethodin was a potent inhibitor of the combined assay, did not inhibit HpkA autophosphorylation, but still led to aggregation of HpkA. These data suggest that ethodin bound to the HpkA kinase and inhibited transfer of the phosphoryl group to DrrA. A peptide corresponding to the phosphorylation site of DrrA appeared to inhibit TCS function by a mechanism similar to that of ethodin, except that autophosphorylation was inhibited at high peptide concentrations. The latter mechanism of inhibition of TCS function is unusual and its analysis demonstrates the utility of these approaches to the kinetic analyses of additional new classes of inhibitors of TCS function.

Probing sesquiterpene hydroxylase activities in a coupled assay with terpene synthases

2003 ◽  
Vol 409 (2) ◽  
pp. 385-394 ◽  
Author(s):  
Bryan T Greenhagen ◽  
Paul Griggs ◽  
Shunji Takahashi ◽  
Lyle Ralston ◽  
Joe Chappell
Keyword(s):  
Terpene Synthases ◽  
Coupled Assay
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