Mutagenesis and biochemical analysis of recombinant yeast prenyltransferases.
The use of the S. cerevisiae protein prenyltransferases as a model system for general prenyltransferase study is justified by the similarity of mechanism, substrate specificity, and evolutionarily conserved substrates with the mammalian prenyltransferases. Genetic identification of potential structural genes involved in prenyltransferase activity can be easily confirmed with biochemical assays using recombinant enzyme reconstitution. Yeast FTase and GGTase I produced in E. coli are indistinguishable from the native proteins and can be studied without interference from contaminating cellular protein prenyltransferases. Structure-function analysis of the yeast prenyltransferase subunits is also simplified by the rapidity with which mutant enzymes can be analyzed in E. coli and their biological activity characterized in yeast defective for the particular subunit gene.
Caplin
B E
BE
Department of Medicine, Indiana University, Indianapolis 46202, USA.
Marshall
M S
MS
D019883:Alkyl and Aryl Transferases; D000595:Amino Acid Sequence; D001483:Base Sequence; D002852:Chromatography, Ion Exchange / Q000379:methods; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology; D051231:Farnesyltranstransferase; D005800:Genes, Fungal; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D011499:Protein Processing, Post-Translational; D011865:Radioisotope Dilution Technique; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D012717:Sesquiterpenes; D013045:Species Specificity; D013379:Substrate Specificity; D014166:Transferases / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014316:Tritium
D017931:DNA Primers; D007202:Indicators and Reagents; D011106:Polyisoprenyl Phosphates; D011994:Recombinant Proteins; D012717:Sesquiterpenes; D014316:Tritium; C004808:farnesyl pyrophosphate; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C083291:geranylgeranyltransferase type-I; D051231:Farnesyltranstransferase; C002963:geranylgeranyl pyrophosphate