Linked Life Data

21455275

Source:http://linkedlifedata.com/resource/pubmed/id/21455275

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2011-4-1
pubmed:abstractText
Nucleoside transporters (NTs) are integral membrane proteins necessary for the cellular entry of nucleoside analog drugs used in chemotherapeutic treatment of conditions such as cancer and viral or parasitic infections. NTs are also the targets of certain drugs used in the treatment of various cardiovascular conditions. Because of the importance of NTs in drug uptake, determination of the three-dimensional structure of these proteins, particularly hENT1, has the potential to improve these treatments through structure-based design of more specifically targeted and transported drugs. In this paper, we use NMR spectroscopy to investigate the structure of the large intracellular loop between transmembrane domains 6 and 7 and we also describe a method for the successful overexpression of full-length hENT1 in a bacterial system. Recombinant tandem histidine-affinity (HAT) and 3×FLAG tagged hENT1 was overexpressed in E. coli, affinity purified, and functionally characterized by nitrobenzylthioinosine (NBTI) binding. Anti-3×FLAG immunodetection confirmed the expression of N-HAT-3×FLAG-hENT1, while increased NBTI binding (3.2-fold compared with controls) confirmed the conformational integrity of the recombinant hENT1 within the bacterial inner membrane. Yields of recombinant hENT1 using this approach were ~15 µg/L of bacterial culture and this approach provides a basis for large-scale production of protein for a variety of purposes.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
1208-6002
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
89
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
246-55
pubmed:meshHeading
pubmed:year
2011
pubmed:articleTitle
Analysis of recombinant tagged equilibrative nucleoside transporter 1 (ENT1) expressed in E. coli.
pubmed:affiliation
Department of Biology, York University, Toronto, ON, Canada.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't