Burkitt lymphoma Daudi cells contain two distinct farnesyltransferases with different divalent cation requirements.
Farnesylation is a lipid posttranslational modification required for the biological function of several signaling proteins including the Ras oncoprotein where this modification is required for malignant transformation. Here we report the identification of two distinct farnesyltransferases (FTases) in Burkitt lymphoma Daudi cells. Separation of Daudi cell cytosolic fractions by ion exchange chromatography resulted in two peaks (FTases I and II) that, on gel filtration, show molecular masses of 90 000 and 250 000 Da, respectively. Immunoblotting experiments showed that FTase I is composed of an alpha/beta-heterodimer of about 50 000 Da each. FTase II contained a beta-subunit that is immunologically indistinguishable from the beta-subunit of FTase I and the previously reported human and rat brain FTase but contained an alpha-subunit that reacted poorly with a rat brain anti-alpha-antibody. As in rat brain FTase, Daudi FTases I and II both required magnesium for enzymatic activity. However, their zinc requirements differed. In the absence of Zn2+, FTase I had little activity (10%) whereas FTase II had 30% of its maximum activity (maximum activity obtained in the presence of Zn2+). Furthermore, whereas both FTases I and II were potently inhibited by KB-Ras C-terminal Cys-Val-Ile-Met tetrapeptide mimics, only FTase I but not FTase II required zinc for peptide binding and inhibition.
Vogt
A
A
Department of Pharmacology, School of Medicine, University of Pittsburgh, Pennsylvania 15261, USA.
Sun
J
J
Qian
Y
Y
Tan-Chiu
E
E
Hamilton
A D
AD
Sebti
S M
SM
D019883:Alkyl and Aryl Transferases; D000818:Animals; D001921:Brain / Q000201:enzymology; D002051:Burkitt Lymphoma / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes / Q000187:drug effects / Q000302:isolation & purification*; D008297:Male; D008970:Molecular Weight; D014066:Palatine Tonsil / Q000201:enzymology; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D011487:Protein Conformation; D017368:Protein Prenylation; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012717:Sesquiterpenes; D014166:Transferases / Q000187:drug effects / Q000302:isolation & purification*; D014407:Tumor Cells, Cultured; D015032:Zinc / Q000494:pharmacology
D007527:Isoenzymes; D011106:Polyisoprenyl Phosphates; D012717:Sesquiterpenes; C004808:farnesyl pyrophosphate; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C064911:p21(ras) farnesyl-protein transferase; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D015032:Zinc