mRNA capping enzyme. Isolation and characterization of the gene encoding mRNA guanylytransferase subunit from Saccharomyces cerevisiae.
The highly purified yeast mRNA capping enzyme is composed of two separate chains of 52 (alpha) and 80 kDa (beta), responsible for the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase, respectively (Itoh, N., Yamada, H., Kaziro, Y., and Mizumoto, K. (1987) J. Biol. Chem. 262, 1989-1995). The gene encoding the mRNA guanylyltransferase subunit (alpha subunit), CEG1, has been isolated by immunological screening of a yeast genomic expression library in lambda gt11 with polyclonal antibodies directed against purified yeast capping enzyme. The identity of CEG1 was confirmed by epitope selection and by expressing the gene in Escherichia coli to give a catalytically active mRNA guanylyltransferase. The gene is present in one copy per haploid genome, and encodes a polypeptide of 459 amino acid residues. From its primary structure as well as its mRNA size, it was concluded that the alpha and the beta subunits of yeast mRNA capping enzyme are encoded by two separate genes, not as a fused protein. CEG1 is located on the chromosome VII by a pulse-field gel electrophoresis. Gene disruption experiment indicated that CEG1 is essential for the growth of yeast. We have also found another open reading frame (ORF2) which lies in close proximity to CEG1 in our clones and encodes a 450 amino acid-polypeptide of yet unknown function.
Shibagaki
Y
Y
Institute of Medical Science, University of Tokyo, Japan.
Itoh
N
N
Yamada
H
H
Nagata
S
S
Mizumoto
K
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005800:Genes, Fungal; D015698:Genomic Library; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000235:genetics*; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics*; D000098556:mRNA Guanylyltransferases
D004271:DNA, Fungal; D046911:Macromolecular Substances; D012333:RNA, Messenger; D009713:Nucleotidyltransferases; D000098556:mRNA Guanylyltransferases