Literature summary for 3.6.1.74 extracted from

Yamada-Okabe, T.; Doi, R.; Shimmi, O.; Arisawa, M.; Yamada-Okabe, H.
Isolation and characterization of a human cDNA for mRNA 5'-capping enzyme (1998), Nucleic Acids Res., 26, 1700-1706 .

Cloned(Commentary)

Cloned (Comment)	Organism
three highly related cDNAs, HCE1 (human mRNA capping enzyme 1), HCE1A, and HCE1B, are cloned from a HeLa cDNA library. DNA and amino acid sequence determination and analysis, sequence comparisons. The HCE1 cDNA has the longest ORF, which can encode a 69 kDa protein. A short region of 69 bp in the 34-half of the HCE1 ORF is missing in HCE1A and HCE1B, and, additionally, HCE1B has an early translation termination signal, which suggests that the latter two cDNAs represent alternatively spliced product. Semiquantitative RT-PCR enzyme expression analysis. Recombinant expression of GST-tagged enzyme variants in Escherichia coli. When expressed from ADH1 promoter, HCE1, but not HCE1A and HCE1B, functionally complements Saccharomyces cerevisiae CEG1 and CET1, the genes for GTase and TPase in yeast, respectively	Homo sapiens

Cloned (Comment)

Organism

three highly related cDNAs, HCE1 (human mRNA capping enzyme 1), HCE1A, and HCE1B, are cloned from a HeLa cDNA library. DNA and amino acid sequence determination and analysis, sequence comparisons. The HCE1 cDNA has the longest ORF, which can encode a 69 kDa protein. A short region of 69 bp in the 34-half of the HCE1 ORF is missing in HCE1A and HCE1B, and, additionally, HCE1B has an early translation termination signal, which suggests that the latter two cDNAs represent alternatively spliced product. Semiquantitative RT-PCR enzyme expression analysis. Recombinant expression of GST-tagged enzyme variants in Escherichia coli. When expressed from ADH1 promoter, HCE1, but not HCE1A and HCE1B, functionally complements Saccharomyces cerevisiae CEG1 and CET1, the genes for GTase and TPase in yeast, respectively

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
E345A	site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants	Homo sapiens
K294A	site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants	Homo sapiens
K458A	site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants	Homo sapiens
K460A	site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants	Homo sapiens
R299A	site-directed mutagenesis, the mutant cannot complement Saccharomyces cerevisiae CEG1 deficient mutants	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
a 5'-triphospho-[mRNA] + H2O	Homo sapiens	-	a 5'-diphospho-[mRNA] + phosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	O60942	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant GST-tagged enzyme variants from Escherichia coli by glutathione affinity separation	Homo sapiens

Source Tissue

Source Tissue	Comment	Organism	Textmining
HeLa cell	-	Homo sapiens	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
a 5'-triphospho-[mRNA] + H2O	-	Homo sapiens	a 5'-diphospho-[mRNA] + phosphate	-	?
a 5'-triphospho-[mRNA] + H2O	[gamma-32P]ATP-terminated RNA	Homo sapiens	a 5'-diphospho-[mRNA] + phosphate	-	?

Subunits

Subunits	Comment	Organism
?	x * 69000, about, HCE1, sequence calculation	Homo sapiens

Synonyms

Synonyms	Comment	Organism
HCE1	-	Homo sapiens
HCE1A	-	Homo sapiens
HCE1B	-	Homo sapiens
Hce1p	-	Homo sapiens
human mRNA capping enzyme 1	-	Homo sapiens
mRNA 5'-capping enzyme	-	Homo sapiens
Tpase	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.9	-	assay at	Homo sapiens

General Information

General Information	Comment	Organism
physiological function	enzyme variant Hce1p displays both mRNA 5'-triphosphatase (TPase) and mRNA 5'-guanylyltransferase (GTase, EC 2.7.7.50) activities, and it forms a cap structure at the 5'-triphosphate end of RNA, demonstrating that it indeed specifies an active mRNA 5'-capping enzyme. The recombinant proteins derived from HCE1A and HCE1B possess only TPase activity. When expressed from ADH1 promoter, HCE1, but not HCE1A and HCE1B, complements Saccharomyces cerevisiae CEG1 and CET1, the genes for GTase and TPase in yeast, respectively. These results demonstrate that the N-terminal part of Hce1p is responsible for TPase activity and the C-terminal part is essential for GTase activity. In addition, the human TPase domain cannot functionally substitute for the yeast enzyme in vivo	Homo sapiens