The modified base J is the target for a novel DNA-binding protein in kinetoplastid protozoans.
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.
Cross
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M
Division of Molecular Biology and Centre of Biomedical Genetics, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands.
Kieft
R
R
Sabatini
R
R
Wilm
M
M
de Kort
M
M
van der Marel
G A
GA
van Boom
J H
JH
van Leeuwen
F
F
Borst
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D016831:Crithidia fasciculata / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020868:Gene Silencing; D005960:Glucosides / Q000378:metabolism*; D016830:Kinetoplastida / Q000235:genetics / Q000378:metabolism*; D007891:Leishmania / Q000235:genetics; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014346:Trypanosoma brucei brucei / Q000235:genetics; D014498:Uracil / Q000031:analogs & derivatives* / Q000378:metabolism; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000235:genetics*
D016054:DNA, Protozoan; D004268:DNA-Binding Proteins; D005960:Glucosides; C402790:J-specific DNA-binding protein, protozoa; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D014643:Variant Surface Glycoproteins, Trypanosoma; C084880:5-((glucopyranosyloxy)methyl)uracil; D014498:Uracil