In Vitro Reconstitution and Kinetic Characterization of the Essential Condensation Enzymes Involved in Prodiginine and Tambjamine Biosynthesis

10.26434/chemrxiv.8001410.v1 ◽

2019 ◽

Author(s):

Katherine J. Picott ◽

Julie A. Deichert ◽

Ella M. deKemp ◽

Avena Ross

Keyword(s):

Substrate Inhibition ◽

Kinetic Characterization ◽

Reaction Sequence ◽

Ping Pong ◽

In Vitro Reconstitution

We have reconstituted<i> </i>the <i>in vitro</i> activity of the purified condensation enzymes PigC and TamQ, from prodiginine and tambjamine pathways respectively. Upon analysis and comparison of their kinetic profiles, a ping-pong reaction sequence with substrate inhibition was identified for each condensation enzyme.

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Membrane-Bound Electron Transfer Chain of the Thermohalophilic BacteriumRhodothermus marinus: Characterization of the Iron−Sulfur Centers from the Dehydrogenases and Investigation of the High-Potential Iron−Sulfur Protein Function by in Vitro Reconstitution of the Respiratory Chain†

Biochemistry ◽

10.1021/bi981807v ◽

1999 ◽

Vol 38 (4) ◽

pp. 1276-1283 ◽

Cited By ~ 29

Author(s):

Manuela M. Pereira ◽

João N. Carita ◽

Miguel Teixeira

Keyword(s):

Protein Function ◽

Iron Sulfur ◽

In Vitro Reconstitution ◽

Sulfur Protein ◽

Membrane Bound ◽

Iron Sulfur Protein ◽

Transfer Chain ◽

Bound Electron

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Effect of Substrate Inhibition and Cooperativity on the Electrochemical Responses of Glucose Dehydrogenase. Kinetic Characterization of Wild and Mutant Types

Journal of the American Chemical Society ◽

10.1021/ja204637d ◽

2011 ◽

Vol 133 (32) ◽

pp. 12801-12809 ◽

Cited By ~ 18

Author(s):

Fabien Durand ◽

Benoît Limoges ◽

Nicolas Mano ◽

François Mavré ◽

Rebeca Miranda-Castro ◽

...

Keyword(s):

Glucose Dehydrogenase ◽

Substrate Inhibition ◽

Kinetic Characterization

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In Vitro Kinetic Characterization of Transporter-Mediated Permeability

Transporters in Drug Development - AAPS Advances in the Pharmaceutical Sciences Series ◽

10.1007/978-1-4614-8229-1_2 ◽

2013 ◽

pp. 23-35

Author(s):

Bente Steffansen ◽

Anne Sophie Grandvuinet

Keyword(s):

Kinetic Characterization

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Kinetic characterization of a cocaine hydrolase engineered from mouse butyrylcholinesterase

Biochemical Journal ◽

10.1042/bj20141266 ◽

2015 ◽

Vol 466 (2) ◽

pp. 243-251 ◽

Cited By ~ 12

Author(s):

Xiabin Chen ◽

Xiaoqin Huang ◽

Liyi Geng ◽

Liu Xue ◽

Shurong Hou ◽

...

Keyword(s):

Amino Acid ◽

Substrate Inhibition ◽

Kinetic Characterization ◽

Substrate Activation ◽

Amino Acid Mutations ◽

Human Butyrylcholinesterase

Mouse (mBChE) and human butyrylcholinesterase (hBChE)-based cocaine hydrolases (mCocH and hCocH) have remarkably different catalytic efficiencies against cocaine, but with little differences in catalytic efficiencies against acetylcholine (ACh) and butyrylthiocholine (BTC). The amino-acid mutations have remarkably converted substrate activation of the enzymes into substrate inhibition.

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Kinetic characterization of inhibitors of histone deacetylases (HDACs) and sirtuins

10.21203/rs.2.13042/v1 ◽

2019 ◽

Cited By ~ 1

Author(s):

Carlos Moreno-Yruela ◽

Andreas Stahl Madsen ◽

Christian A. Olsen

Keyword(s):

Histone Deacetylases ◽

Hdac Inhibitors ◽

Kinetic Properties ◽

Kinetic Characterization ◽

Class Iii ◽

Vitro Assay ◽

Substrate Conversion ◽

Fluorogenic Peptide Substrate

Abstract Histone deacetylase (HDAC) inhibitors are employed for the treatment of lymphoma and are under development against multiple other types of cancer and neurodegenerative diseases. Here, we describe a robust and uncomplicated in vitro assay for HDAC inhibitor kinetic profiling. Enzyme and fluorogenic peptide substrate are incubated together with a small amount of protease “assay developer”, which enables continuous recording of substrate conversion under steady-state conditions. Assay progression curves upon addition of an inhibitors at varying concentrations permit determination of kinetic constants and overall inhibitor potency. This assay helped provide new insight into the kinetic properties of known HDAC inhibitors as well as the kinetic characterization of both inhibitors and substrates of sirtuin enzymes, which are class III HDACs involved in metabolic control and oncogene regulation.

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