The modified base J is the target for a novel DNA-binding protein in kinetoplastid protozoans |
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| Authors: | Cross M Kieft R Sabatini R Wilm M de Kort M van der Marel G A van Boom J H van Leeuwen F Borst P |
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| Affiliation: | Division of Molecular Biology and Centre of Biomedical Genetics, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands. |
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| Abstract: | DNA from Kinetoplastida contains the unusual modified base beta-D-glucosyl(hydroxymethyl)uracil, called J. Base J is found predominantly in repetitive DNA and correlates with epigenetic silencing of telomeric variant surface glycoprotein genes in Trypanosoma brucei. We have now identified a protein in nuclear extracts of bloodstream stage T.brucei that binds specifically to J-containing duplex DNA. J-specific DNA binding was also observed with extracts from the kinetoplastids Crithidia fasciculata and Leishmania tarentolae. We purified the 90 kDa C.fasciculata J-binding protein 50 000-fold and cloned the corresponding gene from C.fasciculata, T.brucei and L.tarentolae. Recombinant proteins expressed in Escherichia coli demonstrated J-specific DNA binding. The J-binding proteins show 43-63% identity and are unlike any known protein. The discovery of a J-binding protein suggests that J, like methylated cytosine in higher eukaryotes, functions via a protein intermediate. |
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