Purification and molecular cloning of a novel essential component of the apolipoprotein B mRNA editing enzyme-complex.
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Purification and molecular cloning of a novel essential component of the apolipoprotein B mRNA editing enzyme-complex. / Lellek, Heinrich; Kirsten, R; Diehl, I; Apostel, F; Buck, F; Greeve, J.
In: J BIOL CHEM, Vol. 275, No. 26, 26, 2000, p. 19848-19856.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - Purification and molecular cloning of a novel essential component of the apolipoprotein B mRNA editing enzyme-complex.
AU - Lellek, Heinrich
AU - Kirsten, R
AU - Diehl, I
AU - Apostel, F
AU - Buck, F
AU - Greeve, J
PY - 2000
Y1 - 2000
N2 - Editing of apolipoprotein B (apoB) mRNA requires the catalytic component APOBEC-1 together with "auxiliary" proteins that have not been conclusively characterized so far. Here we report the purification of these additional components of the apoB mRNA editing enzyme-complex from rat liver and the cDNA cloning of the novel APOBEC-1-stimulating protein (ASP). Two proteins copurified into the final active fraction and were characterized by peptide sequencing and mass spectrometry: KSRP, a 75-kDa protein originally described as a splicing regulating factor, and ASP, a hitherto unknown 65-kDa protein. Separation of these two proteins resulted in a reduction of APOBEC-1-stimulating activity. ASP represents a novel type of RNA-binding protein and contains three single-stranded RNA-binding domains in the amino-terminal half and a putative double-stranded RNA-binding domain at the carboxyl terminus. Purified recombinant glutathione S-transferase (GST)-ASP, but not recombinant GST-KSRP, stimulated recombinant GST-APOBEC-1 to edit apoB RNA in vitro. These data demonstrate that ASP is the second essential component of the apoB mRNA editing enzyme-complex. In rat liver, ASP is apparently associated with KSRP, which may confer stability to the editing enzyme-complex with its substrate apoB RNA serving as an additional auxiliary component.
AB - Editing of apolipoprotein B (apoB) mRNA requires the catalytic component APOBEC-1 together with "auxiliary" proteins that have not been conclusively characterized so far. Here we report the purification of these additional components of the apoB mRNA editing enzyme-complex from rat liver and the cDNA cloning of the novel APOBEC-1-stimulating protein (ASP). Two proteins copurified into the final active fraction and were characterized by peptide sequencing and mass spectrometry: KSRP, a 75-kDa protein originally described as a splicing regulating factor, and ASP, a hitherto unknown 65-kDa protein. Separation of these two proteins resulted in a reduction of APOBEC-1-stimulating activity. ASP represents a novel type of RNA-binding protein and contains three single-stranded RNA-binding domains in the amino-terminal half and a putative double-stranded RNA-binding domain at the carboxyl terminus. Purified recombinant glutathione S-transferase (GST)-ASP, but not recombinant GST-KSRP, stimulated recombinant GST-APOBEC-1 to edit apoB RNA in vitro. These data demonstrate that ASP is the second essential component of the apoB mRNA editing enzyme-complex. In rat liver, ASP is apparently associated with KSRP, which may confer stability to the editing enzyme-complex with its substrate apoB RNA serving as an additional auxiliary component.
M3 - SCORING: Zeitschriftenaufsatz
VL - 275
SP - 19848
EP - 19856
JO - J BIOL CHEM
JF - J BIOL CHEM
SN - 0021-9258
IS - 26
M1 - 26
ER -
