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In vitro Deneddylation Assay

Authors: Anna M. Köhler
Anna M. KöhlerAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
Bio-protocol author page: a2960
, Cindy Meister
Cindy MeisterAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
Bio-protocol author page: a2961
 and Gerhard H. Braus
Gerhard H. BrausAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
For correspondence: gbraus@gwdg.de
Bio-protocol author page: a2962
,  date: 3/20/2016, 1634 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1756.

[Abstract] Nedd8 is a small ubiquitin-like protein (9 kDa) covalently attached to a conserved lysine residue of a cullin protein which is part of cullin-RING ligases (CRLs). CRLs are major E3 ligases important for protein ubiquitination in the ubiquitin-proteasome pathway (UPP). The activity of CRLs is regulated ...

Purification of the GfsA-3x FLAG Protein Expressed in Aspergillus nidulans

Authors: Takuji Oka
Takuji OkaAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
For correspondence: oka@bio.sojo-u.ac.jp
Bio-protocol author page: a1613
, Yukako Katafuchi
Yukako KatafuchiAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1614
, Kohsai Fukuda
Kohsai FukudaAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1615
, Keisuke Ekino
Keisuke EkinoAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1616
, Masatoshi Goto
Masatoshi GotoAffiliation: Department of Bioscience and Biotechnology, Kyushu University, Fukuoka City, Japan
Bio-protocol author page: a1617
 and Yoshiyuki Nomura
Yoshiyuki NomuraAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1618
,  date: 9/5/2014, 2647 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1222.

[Abstract] GfsA is a fungal β-galactofuranosyltransferase involved in the biosynthesis of O-glycan. To investigate the enzymatic functions of GfsA, we attempted to obtain a recombinant protein of this enzyme from two heterologous host organisms. However, GfsA could not be expressed as a recombinant protein in ...

Transport Assays in Aspergillus nidulans

Authors: Emilia Krypotou
Emilia Krypotou Affiliation: Faculty of Biology, University of Athens, Athens, Greece
Bio-protocol author page: a1008
 and George Diallinas
George DiallinasAffiliation: Faculty of Biology, University of Athens, Athens, Greece
For correspondence: diallina@biol.uoa.gr
Bio-protocol author page: a1009
,  date: 11/20/2013, 2705 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.971.

[Abstract] Transport assays allow the direct kinetic analysis of a specific transporter by measuring apparent Km and Vmax values, and permit the characterization of substrate specificity profiles through competition assays. In this protocol, we describe a rapid and easy method for performing uptake assays in the ...

Western Analysis of Histone Modifications (Aspergillus nidulans)

Authors: Alexandra Soukup
Alexandra SoukupAffiliation: Department of Genetics, University of Wisconsin, Madison, USA
Bio-protocol author page: a532
 and Nancy P. Keller
Nancy P. KellerAffiliation: Department of Bacteriology and Department of Medical Microbiology and Immunology, University of Wisconsin, Madison, USA
For correspondence: npkeller@wisc.edu
Bio-protocol author page: a319
,  date: 4/5/2013, 4937 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.424.

[Abstract] Western blotting allows for the specific detection of proteins and/or modifications of proteins by an antibody of interest. This protocol utilizes a crude nuclei extraction protocol for Aspergillus nidulans to enrich for histones and other nuclear proteins prior to gel electrophoresis. Post translational ...
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Western Analysis of Histone Modifications (Aspergillus nidulans)

Authors: Alexandra Soukup
Alexandra SoukupAffiliation: Department of Genetics, University of Wisconsin, Madison, USA
Bio-protocol author page: a532
 and Nancy P. Keller
Nancy P. KellerAffiliation: Department of Bacteriology and Department of Medical Microbiology and Immunology, University of Wisconsin, Madison, USA
For correspondence: npkeller@wisc.edu
Bio-protocol author page: a319
,  date: 4/5/2013, 4937 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.424.

[Abstract] Western blotting allows for the specific detection of proteins and/or modifications of proteins by an antibody of interest. This protocol utilizes a crude nuclei extraction protocol for Aspergillus nidulans to enrich for histones and other nuclear proteins prior to gel electrophoresis. Post translational ...

Transport Assays in Aspergillus nidulans

Authors: Emilia Krypotou
Emilia Krypotou Affiliation: Faculty of Biology, University of Athens, Athens, Greece
Bio-protocol author page: a1008
 and George Diallinas
George DiallinasAffiliation: Faculty of Biology, University of Athens, Athens, Greece
For correspondence: diallina@biol.uoa.gr
Bio-protocol author page: a1009
,  date: 11/20/2013, 2705 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.971.

[Abstract] Transport assays allow the direct kinetic analysis of a specific transporter by measuring apparent Km and Vmax values, and permit the characterization of substrate specificity profiles through competition assays. In this protocol, we describe a rapid and easy method for performing uptake assays in the ...

Purification of the GfsA-3x FLAG Protein Expressed in Aspergillus nidulans

Authors: Takuji Oka
Takuji OkaAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
For correspondence: oka@bio.sojo-u.ac.jp
Bio-protocol author page: a1613
, Yukako Katafuchi
Yukako KatafuchiAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1614
, Kohsai Fukuda
Kohsai FukudaAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1615
, Keisuke Ekino
Keisuke EkinoAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1616
, Masatoshi Goto
Masatoshi GotoAffiliation: Department of Bioscience and Biotechnology, Kyushu University, Fukuoka City, Japan
Bio-protocol author page: a1617
 and Yoshiyuki Nomura
Yoshiyuki NomuraAffiliation: Department of Applied Microbial Technology, Sojo University, Kumamoto City, Japan
Bio-protocol author page: a1618
,  date: 9/5/2014, 2647 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1222.

[Abstract] GfsA is a fungal β-galactofuranosyltransferase involved in the biosynthesis of O-glycan. To investigate the enzymatic functions of GfsA, we attempted to obtain a recombinant protein of this enzyme from two heterologous host organisms. However, GfsA could not be expressed as a recombinant protein in ...

In vitro Deneddylation Assay

Authors: Anna M. Köhler
Anna M. KöhlerAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
Bio-protocol author page: a2960
, Cindy Meister
Cindy MeisterAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
Bio-protocol author page: a2961
 and Gerhard H. Braus
Gerhard H. BrausAffiliation: Abteilung für Molekulare Mikrobiologie und Genetik, Institut für Mikrobiologie und Genetik und Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Göttingen, Germany
For correspondence: gbraus@gwdg.de
Bio-protocol author page: a2962
,  date: 3/20/2016, 1634 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1756.

[Abstract] Nedd8 is a small ubiquitin-like protein (9 kDa) covalently attached to a conserved lysine residue of a cullin protein which is part of cullin-RING ligases (CRLs). CRLs are major E3 ligases important for protein ubiquitination in the ubiquitin-proteasome pathway (UPP). The activity of CRLs is regulated ...
1 
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