MultiDsk: a ubiquitin-specific affinity resin
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MultiDsk : a ubiquitin-specific affinity resin. / Wilson, Marcus D; Saponaro, Marco; Leidl, Mathias A; Svejstrup, Jesper Q.
I: PLOS ONE, Bind 7, Nr. 10, 2012, s. e46398.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - MultiDsk
T2 - a ubiquitin-specific affinity resin
AU - Wilson, Marcus D
AU - Saponaro, Marco
AU - Leidl, Mathias A
AU - Svejstrup, Jesper Q
PY - 2012
Y1 - 2012
N2 - Ubiquitylation is a highly diverse and complex post-translational modification for the regulation of protein function and stability. Studies of ubiquitylation have, however, been hampered by its rapid reversal in cell extracts, for example through the action of de-ubiquitylating enzymes (DUBs). Here we describe a novel ubiquitin-binding protein reagent, MultiDsk, composed of an array of five UBA domains from the yeast ubiquitin-binding protein Dsk2, fused to GST. MultiDsk binds ubiquitylated substrates with unprecedented avidity, and can be used as both an affinity resin to study protein ubiquitylation, and to effectively protect ubiquitylated proteins from the action of DUBs and the proteasome in crude cell extracts. We use the resin to show that the Def1 protein becomes ubiquitylated in response to DNA damage, and to isolate ubiquitylated forms of RNA polymerase II.
AB - Ubiquitylation is a highly diverse and complex post-translational modification for the regulation of protein function and stability. Studies of ubiquitylation have, however, been hampered by its rapid reversal in cell extracts, for example through the action of de-ubiquitylating enzymes (DUBs). Here we describe a novel ubiquitin-binding protein reagent, MultiDsk, composed of an array of five UBA domains from the yeast ubiquitin-binding protein Dsk2, fused to GST. MultiDsk binds ubiquitylated substrates with unprecedented avidity, and can be used as both an affinity resin to study protein ubiquitylation, and to effectively protect ubiquitylated proteins from the action of DUBs and the proteasome in crude cell extracts. We use the resin to show that the Def1 protein becomes ubiquitylated in response to DNA damage, and to isolate ubiquitylated forms of RNA polymerase II.
KW - Amino Acid Sequence
KW - DNA Damage
KW - Humans
KW - Molecular Sequence Data
KW - RNA Polymerase II
KW - Ubiquitin
KW - Ubiquitination
U2 - 10.1371/journal.pone.0046398
DO - 10.1371/journal.pone.0046398
M3 - Journal article
C2 - 23056298
VL - 7
SP - e46398
JO - PLoS ONE
JF - PLoS ONE
SN - 1932-6203
IS - 10
ER -
ID: 47713281