Microbiology

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    Protocols in Current Issue
    Analysis of Gram-negative Bacteria Peptidoglycan by Ultra-performance Liquid Chromatography
    Authors:  Laura Alvarez, Baptiste Cordier, Sven van Teeffelen and Felipe Cava, date: 10/05/2020, view: 288, Q&A: 0
    [Abstract] Bacteria are surrounded by a protective peptidoglycan cell wall. Provided that this structure and the enzymes involved are the preferred target for our most successful antibiotics, determining its structural and chemical complexity is of the highest interest. Traditionally, high-performance liquid chromatography (HPLC) analyses have been ...
    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: 282, 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 ...
    Determination of the Cellular Ion Concentration in Saccharomyces cerevisiae Using ICP-AES
    Authors:  Louise Thines, Anne Iserentant and Pierre Morsomme, date: 08/20/2020, view: 712, Q&A: 0
    [Abstract] The yeast Saccharomyces cerevisiae has been perceived over decades as a highly valuable model organism for the investigation of ion homeostasis. Indeed, many of the genes and biological systems that function in yeast ion homeostasis are conserved throughout unicellular eukaryotes to humans. In this context, measurement of the yeast ...
    A SsrA/NIa-based Strategy for Post-Translational Regulation of Protein Levels in Gram-negative Bacteria
    Authors:  Gonzalo Durante-Rodríguez, Belén Calles, Víctor de Lorenzo and Pablo I. Nikel, date: 07/20/2020, view: 605, Q&A: 0
    [Abstract] Strategies to control the levels of key enzymes of bacterial metabolism are commonly based on the manipulation of gene of interest within the target pathway. The development of new protocols towards the manipulation of biochemical processes is still a major challenge in the field of metabolic engineering. On this background, the FENIX (functional ...
    Site-specific DNA Mapping of Protein Binding Orientation Using Azidophenacyl Bromide (APB)
    Authors:  Himasha M. Perera and Michael A. Trakselis, date: 06/20/2020, view: 1110, Q&A: 0
    [Abstract] The orientation of a DNA-binding protein bound on DNA is determinative in directing the assembly of other associated proteins in the complex for enzymatic action. As an example, in a replisome, the orientation of the DNA helicase at the replication fork directs the assembly of the other associated replisome proteins. We have recently determined ...
    Identification of Buffer Conditions for Optimal Thermostability and Solubility of Herpesviral Protein UL37 Using the Thermofluor Assay
    Authors:  Andrea L. Koenigsberg, Jared D. Pitts and Ekaterina E. Heldwein, date: 06/20/2020, view: 723, Q&A: 0
    [Abstract] Structural and biochemical studies of proteins require high amounts of stable, purified proteins. Protein stability often depends on the buffer composition, which includes pH and concentration of salts or other solutes such as glycerol, hence an efficient method for identifying optimal buffer conditions for stability would minimize time and ...
    Differential Fractionation of Erythrocytes Infected by Plasmodium berghei
    Authors:  Bénédicte Gnangnon, Véronique Peucelle and Christine Pierrot, date: 06/05/2020, view: 731, Q&A: 0
    [Abstract] The study of host/pathogen interactions at the cellular level during Plasmodium intra-erythrocytic cycle requires differential extraction techniques aiming to analyze the different compartments of the infected cell. Various protocols have been proposed in the literature to study specific compartments and/or membranes in the infected ...
    A Sensitive Coupled Enzyme Assay for Measuring Kinase and ATPase Kinetics Using ADP-Specific Hexokinase
    Authors:  Ciaran R. McFarlane and James W. Murray, date: 05/05/2020, view: 1167, Q&A: 0
    [Abstract] Kinases and ATPases perform essential biological functions in metabolism and regulation. Activity of these enzymes is commonly measured by coupling ATP consumption to the synthesis of a detectable product. For most assay systems the ATP concentration during the reaction is unknown, compromising the precision of the assay.

    Using the ...
    Viral Double-Stranded RNA Detection by DNase I and Nuclease S1 digestions in Leishmania parasites
    Authors:  Nathalie Isorce and Nicolas Fasel, date: 05/05/2020, view: 815, Q&A: 0
    [Abstract] Many RNA viruses are found in protozoan parasites. They can be responsible for more serious pathology or treatment failure. For the detection of viral double-stranded RNA (dsRNA), sequence-dependent and -independent methods are available, such as quantitative real-time PCR and immunofluorescence, dot blot, ELISA or sequencing. The technique ...
    Quantification of Nitric Oxide and Reactive Oxygen Species in Leishmania-infected J774A.1 Macrophages as a Response to the in vitro Treatment with a Natural Product Compound
    Authors:  Kalliopi Karampetsou, Olga S. Koutsoni and Eleni Dotsika, date: 12/05/2019, view: 1317, Q&A: 0
    [Abstract] Leishmaniasis is a parasitic disease caused by the obligatory intracellular protozoa Leishmania spp. Current therapeutic options are limited and thus, drug discovery against leishmaniasis is very important. Nevertheless, there is a great difficulty to develop therapeutic strategies against the disease because the parasite deploys various ...



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