Linked Life Data

1400305

Source:http://linkedlifedata.com/resource/pubmed/id/1400305

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
28
pubmed:dateCreated
1992-11-16
pubmed:abstractText
A plasmid has been constructed by cloning the complete crtI gene encoding phytoene desaturase from Erwinia uredovora behind the lac Z promoter of pUC18 resulting in a reading frame for the full polypeptide with additional 9 amino acids at the N terminus. This plasmid mediated the overexpression of phytoene desaturase in transformed Escherichia coli. The overexpressed enzyme was sequestrated into inclusion bodies requiring urea treatment for solubilization. Purification to homogeneity was subsequently performed on a DEAE-cellulose column and by SDS-polyacrylamide gel electrophoresis. The purification scheme allowed the isolation of 5.3 mg of homogeneous desaturase protein from 100 ml of E. coli cell suspension. On SDS-polyacrylamide gel electrophoresis an apparent molecular mass of 56.2 kDa was determined. An antiserum raised against phytoene desaturase cross-reacted with the expressed protein and was employed to monitor the isolation steps. Upon removal of urea, desaturase activity was restored. The isolated desaturase catalyzed the conversion of 15-cis-phytoene to trans-lycopene as well as to bisdehydrolycopene. FAD was involved in desaturation, whereas NAD and NADP were inhibitory. This is the first time that a membrane-integrated carotenogenic enzyme has been purified and finally obtained in an active state.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
267
pubmed:geneSymbol
crtI, lacZ
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
19891-5
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
1992
pubmed:articleTitle
Expression in Escherichia coli, purification, and reactivation of the recombinant Erwinia uredovora phytoene desaturase.
pubmed:affiliation
Lehrstuhl für Physiologie und Biochemie der Pflanzen, Universität Konstanz, Germany.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't