Plant Science

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    Protocols in Current Issue
    In vitro Assays to Evaluate Specificity and Affinity in Protein-phospholipid Interactions
    [Abstract]

    Protein–lipid interactions play important roles in many biological processes, including metabolism, signaling, and transport; however, computational and structural analyses often fail to predict such interactions, and determining which lipids participate in these interactions remains challenging. In vitro assays to assess the physical

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    Apoplastic Expression of CARD1-ecto Domain in Nicotiana benthamiana and Purification from the Apoplastic Fluids
    Authors:  Nobuaki Ishihama, Anuphon Laohavisit, Kaori Takizawa and Ken Shirasu, date: 04/20/2022, view: 954, Q&A: 0
    [Abstract]

    The protein expression and purification process is an essential initial step for biochemical analysis of a protein of interest. Traditionally, heterologous protein expression systems (such as E. coli, yeast, insect cells, and cell-free) are employed for plant protein expression, although a plant expression system is often desirable for plant

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    In vitro Auto- and Substrate-Ubiquitination Assays
    Authors:  Hye Lin Park, Han Yong Lee and Gyeong Mee Yoon, date: 04/05/2022, view: 582, Q&A: 0
    [Abstract]

    The precise regulation of the homeostasis of the cellular proteome is critical for the appropriate growth and development of plants. It also allows the plants to respond to various environmental stresses, by modulating their biochemical and physiological aspects in a timely manner. Ubiquitination of cellular proteins is one of the major

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    Activity-based Protein Profiling of Serine Hydrolase Superfamily Enzymes
    Authors:  Achintya Kumar Dolui, Mahadev Latha and Panneerselvam Vijayaraj, date: 03/20/2022, view: 624, Q&A: 0
    [Abstract]

    Activity-based protein profiling (ABPP) is a chemoproteomics platform to assess the functional state of enzymes in complex biological systems. Over the two decades, ABPP has emerged from a gel-based to gel-free platform, for in-depth proteome analysis with enhanced resolution, sensitivity for target detection, and discovery of small molecule

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    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: 888, 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: 1308, 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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    Proteoliposomes for Studying Lipid-protein Interactions in Membranes in vitro
    Author:  Helmut Kirchhoff, date: 10/20/2021, view: 1021, Q&A: 0
    [Abstract]

    Lipids in biomembranes can control the structure and, therefore, the functionality of membrane-embedded protein complexes. Unraveling how the lipid composition determines the mode of operation of membrane proteins provides mechanistic insights into their functionality. We applied a proteoliposome technique for studying how proteins function in

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    Solubilization Method for Isolation of Photosynthetic Mega- and Super-complexes from Conifer Thylakoids
    Authors:  Pushan Bag, Wolfgang P. Schröder, Stefan Jansson and Domenica Farci, date: 09/05/2021, view: 1541, Q&A: 0
    [Abstract]

    Photosynthesis is the main process by which sunlight is harvested and converted into chemical energy and has been a focal point of fundamental research in plant biology for decades. In higher plants, the process takes place in the thylakoid membranes where the two photosystems (PSI and PSII) are located. In the past few decades, the evolution of

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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: 2989, 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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    In vitro Reconstitution Assays of Arabidopsis 20S Proteasome
    Authors:  Yanjun Li, Di Sun, Xingxing Yan, Zhiye Wang and Xiuren Zhang, date: 04/05/2021, view: 3872, Q&A: 0
    [Abstract]

    The majority of cellular proteins are degraded by the 26S proteasome in eukaryotes. However, intrinsically disordered proteins (IDPs), which contain large portions of unstructured regions and are inherently unstable, are degraded via the ubiquitin-independent 20S proteasome. Emerging evidence indicates that plant IDP homeostasis may also be

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