Molecular cloning and characterization of the yeast gene for squalene synthetase.
Squalene synthetase (farnesyl-diphosphate: farnesyl-diphosphate farnesyltransferase, EC 2.5.1.21) is a critical branch point enzyme of isoprenoid biosynthesis that is thought to regulate the flux of isoprene intermediates through the sterol pathway. The structural gene for this enzyme was cloned from the yeast Saccharomyces cerevisiae by functional complementation of a squalene synthetase-deficient erg9 mutant. Identification of this ERG9 clone was confirmed by genetic linkage analysis in yeast and expression of enzyme activity in Escherichia coli. The predicted squalene synthetase polypeptide of 444 amino acids (Mr, 51,753) lacks significant homology to known protein sequences, except within a region that may represent a prenyl diphosphate (substrate) binding site. The ERG9-encoded protein contains a PEST consensus motif (rich in proline, glutamic acid, serine, and threonine) present in many proteins with short cellular half-lives. Modeling of the protein suggests that it contains at least one, and possibly two, membrane-spanning domains. Disruption of the chromosomal squalene synthetase coding region by insertional mutagenesis indicates that ERG9 is a single copy gene that is essential for cell growth in yeast.
Jennings
S M
SM
Department of Cellular Biology, Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, NJ 08543-4000.
Tsay
Y H
YH
Fisch
T M
TM
Robinson
G W
GW
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D015723:Gene Library; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
D011994:Recombinant Proteins; D013186:Farnesyl-Diphosphate Farnesyltransferase