Linked Life Data

10833404

Source:http://linkedlifedata.com/resource/pubmed/id/10833404

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
2000-8-14
pubmed:abstractText
Expression of the membrane-bound form of rabbit cytochrome b(5) in Escherichia coli has been significantly improved through the use of the T7 expression vector pLW01 (A. Bridges, L. Gruenke, Y.-T. Chang, I. Vakser, G. Loew, and L. Waskell, 1998, J. Biol. Chem. 273, 17036-17049) in conjunction with strain C41(DE3) (B. Miroux and J. Walker, 1996, J. Mol. Biol. 260, 289-298). Cell cultures expressing the cytochrome b(5) contained an average of 820 mg/liter of culture and reached peak levels as high as 1100 mg/liter when higher antibiotic concentrations were used. Maximal levels were obtained from cultures when expression was induced with 10 microM IPTG. Approximately 90% of the cytochrome b(5) was expressed as apoprotein which was reconstituted by addition of exogenous heme. The cytochrome b(5) was purified from detergent-solubilized bacterial membranes using anion-exchange chromatography on DEAE-Sepharose followed by size-exclusion chromatography on Superdex-75. Purification of cytochrome b(5) from a 500-ml culture yielded 121 mg of protein which had a specific content of 50 nmol of heme per milligram of protein with an overall recovery of 35%. The final cytochrome b(5) was free of any detectable contaminants when analyzed by SDS-PAGE.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
1046-5928
pubmed:author
pubmed:copyrightInfo
Copyright 2000 Academic Press.
pubmed:issnType
Print
pubmed:volume
19
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
173-8
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
High-level expression in Escherichia coli and purification of the membrane-bound form of cytochrome b(5).
pubmed:affiliation
Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48109-0606, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.