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Faustovirus Capping Enzyme (FCE) catalyzes the addition of N7-methylguanosine cap (m7G) to the 5′ end of triphosphorylated and diphosphorylated transcripts, producing Cap-0 RNA. FCE is a single-subunit enzyme that combines the three activities necessary to produce the Cap-0 structure- triphosphatase, guanylyltransferase, and (guanine-N7)-methyltransferase. Installation of Cap-0 is a key step in eukaryotic mRNA maturation along with 2′-O-methylation at position 1 (Cap-1) and polyadenosine (poly(A)) tailing. The Cap-0 structure also promotes RNA stability and prevents inadvertent activation of innate immune responses triggered by triphosphorylated RNA.
FCE retains significant capping activity at low temperatures and tolerates reaction temperatures up to 55°C. In many cases, 1 µl of FCE (25 units) can cap over 100 µg of RNA in 1 hour at 37°C. GTP and S-adenosylmethionine (SAM) are required for capping activity and are included with the enzyme.
Source: An E. coli strain that carries a plasmid encoding the Faustovirus Capping Enzyme with a C-terminal His-tag.
Reaction Conditions
1X FCE Capping Buffer
Supplement with 0.5 mM GTP and 0.1 mM S-adenosylmethionine (SAM)
Incubate at 37°C
1X FCE Capping Buffer
50 mM Tris-HCl
5 mM KCl
1 mM MgCl2
1 mM DTT
0.02% Poloxamer 188
(pH 8 @ 25.0°C)
Storage Buffer
40 mM Tris-HCl
100 mM NaCl
50 mM Arginine
0.1 mM TCEP
50% Glycerol
pH 8 @ 25°C
Heat Inactivation
70°C for 10 minutes
Addition of EDTA to 5 mM is recommended to avoid RNA hydrolysis.
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| Description |
Faustovirus Capping Enzyme (FCE) catalyzes the addition of N7-methylguanosine cap (m7G) to the 5′ end of triphosphorylated and diphosphorylated transcripts, producing Cap-0 RNA. FCE is a single-subunit enzyme that combines the three activities necessary to produce the Cap-0 structure- triphosphatase, guanylyltransferase, and (guanine-N7)-methyltransferase. Installation of Cap-0 is a key step in eukaryotic mRNA maturation along with 2′-O-methylation at position 1 (Cap-1) and polyadenosine (poly(A)) tailing. The Cap-0 structure also promotes RNA stability and prevents inadvertent activation of innate immune responses triggered by triphosphorylated RNA. Reaction Conditions 1X FCE Capping Buffer 1X FCE Capping Buffer Storage Buffer 40 mM Tris-HCl Heat Inactivation 70°C for 10 minutes Addition of EDTA to 5 mM is recommended to avoid RNA hydrolysis.
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