Activity —Protein —Plant biochemistry —Plant Science

An in vitro Coupled Assay for PEPC with Control of Bicarbonate Concentration

Authors: Nicholas R. Moody, Chatawal Phansopal and James D. Reid, date: 12/20/2021, view: 986, Q&A: 0

[Abstract]

Phosphoenolpyruvate carboxylase (PEPC) catalyzes a critical step in carbon metabolism in plants and bacteria, the irreversible reaction between bicarbonate and phosphoenolpyruvate to produce the C4 compound oxaloacetate. This enzyme is particularly important in the context of C4 photosynthesis, where it is the initial carbon-fixing enzyme. Many

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High Throughput Analyses of Ascorbate-turnover Enzyme Activities in Rice (Oryza sativa L.) Seedlings

Authors: Lin-Bo Wu, Yanru Feng, Frederike Zeibig, Muhammad Shahedul Alam and Michael Frei, date: 10/20/2021, view: 1463, Q&A: 0

[Abstract]

Ascorbate (Vitamin C) fulfills various functions in plant photosynthesis and abiotic stress tolerance. The four key enzymes involved in the ascorbate-turnover pathway are ascorbate peroxidase, ascorbate oxidase, monodehydroascorbate reductase, and dehydroascorbate reductase. Several reports have shown the pivotal roles of these enzymes in plant

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Characterising Plant Deubiquitinases with in vitro Activity-based Labelling and Ubiquitin Chain Disassembly Assays

Authors: Michael J. Skelly and Steven H. Spoel, date: 05/05/2021, view: 3079, Q&A: 0

[Abstract]

Post-translational modification of proteins by ubiquitin is an essential cellular signaling mechanism in all eukaryotes. Ubiquitin is removed from target proteins by a wide range of deubiquitinase (DUB) enzymes with different activities and substrate specificities. Understanding how DUBs function in vitro is a vital first step to uncovering their

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Non-radioactive Assay to Determine Product Profile of Short-chain Isoprenyl Diphosphate Synthases

Authors: Avanish Rai and Dinesh A. Nagegowda, date: 01/05/2021, view: 2140, Q&A: 0

[Abstract] Isoprenoids represent the largest class of metabolites with amazing diversities in structure and function. They are involved in protecting plants against pathogens or herbivores or involved in attracting pollinators. Isoprenoids are derived from geranyl diphosphate (GPP; C₁₀), farnesyl diphosphate (FPP; C₁₅), geranylgeranyl ...

The ATPase Activity of Escherichia coli Expressed AAA⁺-ATPase Protein

Authors: Amita Kaundal, Vemanna S. Ramu and Kirankumar S. Mysore, date: 08/05/2020, view: 2995, Q&A: 0

[Abstract] ATPases are the enzymes that breakdown ATP to ADP and release inorganic phosphate (Pi). Here we provide a detailed protocol to determine the ATPase activity of a recombinant AAA⁺-ATPase protein (GENERAL CONTROL NON-REPRESSIBLE-4 [GCN4]) by spectrophotometric absorption at 360 nm to measure the accumulated inorganic phosphate. In ...

Enzymatic Assays and Enzyme Histochemistry of Tuta absoluta Feeding on Tomato Leaves

Authors: Rim Hamza, José P. Beltrán and Luis A. Cañas, date: 09/05/2018, view: 5843, Q&A: 0

[Abstract] Enzymes play a key role in insect-plant relationships. For a better understanding of these interactions, we analyzed Tuta absoluta digestive enzymes. Here, we describe a detailed protocol for the detection of trypsin and papain-like enzymes in Tuta absoluta larvae by enzyme histochemistry. This assay uses frozen and unfixed ...

In vitro Nitrate Reductase Activity Assay from Arabidopsis Crude Extracts

Authors: Joo Yong Kim and Hak Soo Seo, date: 04/05/2018, view: 8546, Q&A: 0

[Abstract] Nitrate reductase (NR) reduces the major plant nitrogen source, NO₃^-, into NO₂^-. NR activity can be measured by its final product, nitrite through its absorbance under optimized condition. Here, we present a detailed protocol for measuring relative enzyme activity of NR from Arabidopsis crude ...

In vitro RNA-dependent RNA Polymerase Assay Using Arabidopsis RDR6

Authors: Kyungmin Baeg, Yukihide Tomari and Hiro-oki Iwakawa, date: 01/05/2018, view: 8729, Q&A: 0

[Abstract] RNA-dependent RNA polymerases (RdRPs) in eukaryotes convert single-stranded RNAs into double-stranded RNAs, thereby amplifying small interfering RNAs that play crucial roles in the regulation of development, maintenance of genome integrity and antiviral immunity. Here, we describe a method of in vitro RdRP assay using recombinant ...

Determination of H⁺-ATPase Activity in Arabidopsis Guard Cell Protoplasts through H⁺-pumping Measurement and H⁺-ATPase Quantification

Authors: Shota Yamauchi and Ken-ichiro Shimazaki, date: 12/20/2017, view: 7780, Q&A: 0

[Abstract] The opening of stomata in plants in response to blue light is driven by the plasma membrane H⁺-ATPase in guard cells. To evaluate the activation of the H⁺-ATPase in vivo, we can use H⁺-pumping by guard cells in response to blue light and fusicoccin. To do this, it is required to prepare a large amount of ...

Isolation and Detection of the Chlorophyll Catabolite Hydroxylating Activity from Capsicum annuum Chromoplasts

Authors: Mareike Hauenstein and Stefan Hörtensteiner, date: 09/20/2017, view: 7153, Q&A: 0

[Abstract] Hydroxylation of chlorophyll catabolites at the so-called C3² position (Hauenstein et al., 2016) is commonly found in all plant species analyzed to date. Here we describe an in vitro hydroxylation assay using Capsicum annuum chromoplast membranes as a source of the hydroxylating activity, which converts the ...

Plant Science

Evaluation of Urine Proteins by Capillary Electrophoresis

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