Linked Life Data

1446683

Source:http://linkedlifedata.com/resource/pubmed/id/1446683

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1992-12-30
pubmed:abstractText
We have developed a straightforward, two-step procedure to isolate highly purified yeast tubulin that reproducibly assembles into microtubules. The starting extracts are obtained from cells genetically engineered to overproduce both the alpha and beta subunits of tubulin, under control of the galactose promoter, to approximately 10-times wild-type levels. The first step of purification is carried out with the high-speed supernatant of lysed cells loaded onto a DEAE-Sephadex column; after this step the tubulin preparation is approximately 30% pure. In the second step, the tubulin fractions are loaded onto an immunoaffinity column prepared by coupling the anti-(alpha-tubulin) monoclonal antibody YL 1/2 to Sepharose-4B. Following elution with 0.8 M KCl, the tubulin present in the peak is 90% pure. Upon addition of porcine brain microtubule-associated proteins or DEAE-dextran, this tubulin preparation is functionally active for assembly into microtubules, as visualized by electron microscopy on negatively stained samples. Virtually identical microtubule structures are produced in parallel experiments on the assembly of yeast or porcine brain tubulin, with differences observed only at acidic pH values. Overall, this relatively simple procedure provides a useful tool for the production of functional tubulin suitable both for structural studies and for investigations of the assembly process.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
0014-2956
pubmed:author
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
210
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
343-9
pubmed:dateRevised
2007-7-23
pubmed:meshHeading
pubmed:year
1992
pubmed:articleTitle
Purification of assembly-competent tubulin from Saccharomyces cerevisiae.
pubmed:affiliation
Department of Biochemistry, University of Geneva, Switzerland.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't