Linked Life Data

18171622

Source:http://linkedlifedata.com/resource/pubmed/id/18171622

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2008-2-27
pubmed:abstractText
Bacterial over-expression of proteins is a powerful tool to obtain soluble protein amenable to biochemical, biophysical and/or structural characterization. However, it is well established that many recombinant proteins cannot be produced in a soluble form. Several theoretical and empirical methods to improve soluble production have been suggested, although there is to date no universally accepted protocol. This report describes, and quantitatively analyses, a systematic multi-construct approach to obtain soluble protein. Although commonly used in several laboratories, quantitative analyses of the merits of the strategy applied to a larger number of target proteins are missing from the literature. In this study, typically 10 different protein constructs were tested for each targeted domain of nearly 400 human proteins. Overall, soluble expression was obtained for nearly 50% of the human target proteins upon over-expression in Escherichia coli. The chance of obtaining soluble expression was almost doubled using the multi-construct method as compared to more traditional approaches. Soluble protein constructs were subsequently subjected to crystallization trials and the multi-construct approach yielded a more than fourfold increase, from 15 proteins to 65, for the likelihood of obtaining well-diffracting crystals. The results also demonstrate the value of testing multiple constructs in crystallization trials. Finally, a retrospective analysis of gel filtration profiles indicates that these could be used with caution to prioritize protein targets for crystallization trials.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
1096-0279
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
58
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
210-21
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
The use of systematic N- and C-terminal deletions to promote production and structural studies of recombinant proteins.
pubmed:affiliation
Structural Genomics Consortium, Karolinska Institutet, Department of Medical Biophysics and Biochemistry, 17177 Stockholm, Sweden. susanne.graslund@ki.se
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't