Biochemistry

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    Protocols in Current Issue
    A Co-purification Method for Efficient Production and Src Kinase-mediated Phosphorylation of Aplysia Cortactin
    Authors:  Sherlene L. Brown, Yuan Ren, Daniel M. Suter and Seema Mattoo, date: 09/20/2021, view: 1238, Q&A: 0
    [Abstract]

    Cortactin is an actin-binding protein that regulates processes like cell migration, endocytosis, and tumor cell metastasis. Although cortactin is associated with actin-cytoskeletal dynamics in non-neuronal cells and cell-free systems, the exact mechanisms underlying its fundamental roles in neuronal growth cones are not fully explored. Recent

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    Protein Import Assay into Mitochondria Isolated from Human Cells
    Authors:  Lena M. Murschall, Esra Peker, Thomas MacVicar, Thomas Langer and Jan Riemer, date: 06/20/2021, view: 2546, Q&A: 0
    [Abstract]

    Mitochondria are essential organelles containing approximately 1,500 proteins. Only approximately 1% of these proteins are synthesized inside mitochondria, whereas the remaining 99% are synthesized as precursors on cytosolic ribosomes and imported into the organelle. Various tools and techniques to analyze the import process have been developed.

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    Proteomics Profiling of S-sulfurated Proteins in Acinetobacter baumannii
    Authors:  Brenna J. C. Walsh and David P. Giedroc, date: 05/05/2021, view: 2901, Q&A: 0
    [Abstract]

    Hydrogen sulfide (H2S) is emerging as an important modulator in bacterial cytoprotection against the host immune response in infected animals, which may well be attributed to downstream highly oxidized sulfur species, termed reactive sulfur species (RSS), derived from H2S. One mechanism by which H2S/RSS may signal in the cell is through proteome

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    Click Chemistry for Imaging in-situ Protein Palmitoylation during the Asexual Stages of Plasmodium falciparum
    Author:  Mansoor A. Siddiqui, date: 05/05/2021, view: 3298, Q&A: 0
    [Abstract]

    Palmitoylation refers to the modification of the cysteine thiols in proteins by fatty acids, most commonly palmitic acid, through ‘thioester bond’ formation. In vivo, palmitoylation of proteins is catalyzed by palmitoyl acyltransferases (PATs or DHHC-PATs). Palmitoylation has recently emerged as a crucial post-translational

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    In vitro Glutamylation Inhibition of Ubiquitin Modification and Phosphoribosyl-Ubiquitin Ligation Mediated by Legionella pneumophila Effectors
    Authors:  Alan G. Sulpizio, Marena E. Minelli and Yuxin Mao, date: 11/05/2020, view: 1758, Q&A: 0
    [Abstract]

    Glutamylation is a posttranslational modification where the amino group of a free glutamate amino acid is conjugated to the carboxyl group of a glutamate side chain within a target protein. SidJ is a Legionella kinase-like protein that has recently been identified to perform protein polyglutamylation of the Legionella SdeA Phosphoribosyl-Ubiquitin

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    Radioactive Assay of in vitro Glutamylation Activity of the Legionella pneumophila Effector Protein SidJ
    Authors:  Alan G. Sulpizio, Jung-Ho Shin, Marena E. Minelli and Yuxin Mao, date: 10/05/2020, view: 2121, Q&A: 0
    [Abstract] The Legionella effector protein SidJ has recently been identified to perform polyglutamylation on another Legionella effector, SdeA, ablating SdeA’s activity. SidJ is a kinase-like protein that requires the small eukaryotic protein calmodulin to perform glutamylation. Glutamylation is a relatively uncommon type of ...
    A Workflow for Ultra-rapid Analysis of Histone Post-translational Modifications with Direct-injection Mass Spectrometry
    Authors:  Natarajan V. Bhanu, Simone Sidoli and Benjamin A Garcia, date: 09/20/2020, view: 3416, Q&A: 0
    [Abstract] Chromatin modifications, like histone post translational modifications (PTMs), are critical for tuning gene expression and many other aspects of cell phenotype. Liquid chromatography coupled to mass spectrometry (LC-MS) has become the most suitable method to analyze histones and histone PTMs in a large-scale manner. Selected histone PTMs have ...
    Electrophoretic Mobility Shift Assay of in vitro Phosphorylated RNA Polymerase II Carboxyl-terminal Domain Substrates
    Authors:  Joshua E. Mayfield, Seema Irani and Yan Zhang, date: 06/20/2020, view: 2975, Q&A: 0
    [Abstract] Eukaryotic RNA polymerase II transcribes all protein-coding mRNAs and is highly regulated. A key mechanism directing RNA polymerase II and facilitating the co-transcriptional processing of mRNAs is the phosphorylation of its highly repetitive carboxyl-terminal domain (CTD) of its largest subunit, RPB1, at specific residues. A variety of techniques ...
    In vitro Crosslinking Reactions and Substrate Incorporation Assays for The Identification of Transglutaminase-2 Protein Substrates
    Authors:  William L. Willis, Abigail Foster, Caitlin Henry, Lai Chu Wu and Wael Jarjour, date: 06/20/2020, view: 2236, Q&A: 0
    [Abstract] Transglutaminase (TG2) catalyzes protein crosslinking between glutamyl and lysyl residues. Catalytic activity occurs via a transamidation mechanism resulting in the formation of isopeptide bonds. Since TG2-mediated transamidation is of mechanistic importance for a number of biological processes, assays that enable rapid and efficient ...
    Preparation, FPLC Purification and LC-FT-ICR-MS of Proteins
    Authors:  Tyler B. Johnson, Jiri Adamec and Paul Blum, date: 04/05/2020, view: 3208, Q&A: 0
    [Abstract] High magnetic field Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometers provide extremely high mass resolution (resolving power of ~200,000 at 400 m/z) protein detection across a broad mass range, enabling analysis of fine structure of isotopic peak clusters that is missed in other types of mass spectrometers. The protocol ...



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