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Partial purification of a Leishmania tarentolae protein that binds a putative 5S rRNA promoter element
Author(s) -
Okezie Oneybuchi,
Jagus Rosemary,
Pitula Joseph
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1003.15
Subject(s) - promoter , biology , transcription (linguistics) , microbiology and biotechnology , leishmania , rna polymerase iii , rna polymerase ii , rna , gene , ribosomal rna , rna polymerase , genetics , gene expression , parasite hosting , linguistics , philosophy , world wide web , computer science
Kinetoplastids are a group of protozoan parasites that have been intensively studied due to their adverse impact on agriculture and human health. As in higher eukaryotes, 5S rRNAs in kinetoplastids are derived from RNA polymerase III (pol III) transcription. Despite the presence of pol III activity for 5S rRNA transcription characterization of the RNA pol III promoters of kinetoplastids and the transcription factors (TF) binding them remains meager. Thus, we have initiated a biochemical approach towards the identification of TFIII proteins from the model kinetoplastid Leishmania tarentolae. Specifically, we have employed a putative 5S rDNA pol III promoter sequence as the target for TFIII‐specific interaction by gel shift analysis. Our experiments show that this promoter sequence is specifically recognized by at least one protein factor from a L. tarentolae nuclear extract, as demonstrated through competition assays with various 5S rDNA‐specific constructs. In addition, we have employed double stranded DNA affinity chromatography followed by gel shift analysis as an initial step in purification of this putative 5S rDNA gene‐specific binding protein factor.