Linked Life Data

16413259

Source:http://linkedlifedata.com/resource/pubmed/id/16413259

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:dateCreated
2006-1-17
pubmed:abstractText
In order to reconstitute the generation of COPII vesicles from synthetic liposomes, the minimum requirements are the coat components, Sar1p GTPase, Sec23/24p, Sec13/31p, and a nonhydrolyzable GTP analog such as GMP-PNP. However, in the presence of GTP, nucleotide hydrolysis by Sar1p renders the coat insufficiently stable to sustain vesicle budding. Sar1p GTPase activity was activated by the Sec23/24p GTPase-activating protein (GAP), and further accelerated 10-fold by Sec13/31p. In order to study GTP-dependent budding, we introduced the Sar1p guanine nucleotide exchange factor (GEF), Sec12p. We evaluated Sar1p activation by Sec12p and the dynamics of coat assembly and disassembly in the presence of both Sec12p and Sec23/24p. The cytoplasmic domain of Sec12p activated Sar1p with a turnover 10-fold higher than the GAP activity of Sec23p in the presence of Sec13/31p. As a result, the entire COPII coat remains stable in the presence of GTP. Here, we describe methods to purify Sec12p, real-time fluorescence assays to evaluate COPII coat formation, and the relevant kinetic analyses.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
0076-6879
pubmed:author
pubmed:issnType
Print
pubmed:volume
404
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
74-82
pubmed:dateRevised
2009-11-19
pubmed:meshHeading
pubmed:year
2005
pubmed:articleTitle
Purification and functional properties of yeast Sec12 GEF.
pubmed:affiliation
Department of Molecular and Cell Biology, University of California, Berkeley, USA.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't