Linked Life Data

12615322

Source:http://linkedlifedata.com/resource/pubmed/id/12615322

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2003-3-4
pubmed:databankReference
pubmed:abstractText
Plasmodium falciparum iron regulatory-like protein (PfIRPa, accession AJ012289) has homology to a family of iron-responsive element (IRE)-binding proteins (IRPs) found in different species. We have previously demonstrated that erythrocyte P. falciparum PfIRPa binds a mammalian consensus IRE and that the binding activity is regulated by iron status. In the work we now report, we have cloned a C-terminus histidine-tagged PfIRPa and overexpressed it in a bacterial expression system in soluble form capable of binding IREs. To overexpress PfIRPa, we used the T7 promoter-driven vector, pET28a(+), in conjunction with the Rosetta(DE3)pLysS strain of E. coli, which carries extra copies of tRNA genes usually found in organisms such as P. falciparum whose genome is (A+T)-rich. The histidine-tagged recombinant protein (rPfIRPa) in soluble form was partially purified using His-bind resin. We searched the plasmodial database, plasmoDB, to identify sequences capable of forming IRE loops using a specially developed algorithm, and found three plasmodial sequences matching the search criteria. In gel retardation assays, rPfIRPa bound three 32P-labeled putative plasmodial IREs with affinity exceeding the affinity for the mammalian consensus IRE. The binding was concentration-dependent and was not inhibited by heparin, an inhibitor of non-specific binding. Immunodepletion of rPfIRPa resulted in substantial inhibition of the signal intensity in the gel retardation assays and in Western blot-determinations of rPfIRPa protein levels. Endogenous PfIRPa retained all three putative 32P-IREs at the same position on the gel as the recombinant PfIRPa.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Feb
pubmed:issn
0166-6851
pubmed:author
pubmed:copyrightInfo
Copyright 2002 Elsevier Science B.V.
pubmed:issnType
Print
pubmed:volume
126
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
231-8
pubmed:dateRevised
2009-11-19
pubmed:meshHeading
pubmed-meshheading:12615322-Animals, pubmed-meshheading:12615322-Base Sequence, pubmed-meshheading:12615322-Binding Sites, pubmed-meshheading:12615322-DNA Primers, pubmed-meshheading:12615322-Humans, pubmed-meshheading:12615322-Iron-Regulatory Proteins, pubmed-meshheading:12615322-Jurkat Cells, pubmed-meshheading:12615322-Molecular Sequence Data, pubmed-meshheading:12615322-Nucleic Acid Conformation, pubmed-meshheading:12615322-Plasmodium falciparum, pubmed-meshheading:12615322-Protein Binding, pubmed-meshheading:12615322-Protein-Tyrosine Kinases, pubmed-meshheading:12615322-Proto-Oncogene Proteins, pubmed-meshheading:12615322-Protozoan Proteins, pubmed-meshheading:12615322-RNA, Protozoan, pubmed-meshheading:12615322-Recombinant Proteins, pubmed-meshheading:12615322-Wnt Proteins, pubmed-meshheading:12615322-Zebrafish Proteins
pubmed:year
2003
pubmed:articleTitle
Expression of a recombinant IRP-like Plasmodium falciparum protein that specifically binds putative plasmodial IREs.
pubmed:affiliation
Center for Sickle Cell Disease, Howard University, 2121 Georgia Avenue, NW, Washington, DC 20059, USA. mloyevsky@howard.edu
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.