DB code: T00050

RLCP classification 3.133.300000.396 : Transfer
CATH domain 3.90.63.10 : mRNA Capping Enzyme; Chain A, domain 1 Catalytic domain
2.40.50.140 : OB fold (Dihydrolipoamide Acetyltransferase, E2P) Catalytic domain
4.10.87.10 : mRNA Capping Enzyme; Chain Catalytic domain
E.C. 2.7.7.50
CSA
M-CSA
MACiE

CATH domain Related DB codes (homologues)
2.40.50.140 : OB fold (Dihydrolipoamide Acetyltransferase, E2P) M00220 M00186 D00291 D00294 T00254

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq Pfam
Q84424 mRNA-capping enzyme
GTP--RNA guanylyltransferase
mRNA guanylyltransferase
EC 2.7.7.50
NP_048451.1 (Protein)
NC_000852.5 (DNA/RNA sequence)
PF03919 (mRNA_cap_C)
PF01331 (mRNA_cap_enzyme)
[Graphical View]
P78587 mRNA-capping enzyme subunit alpha
GTP--RNA guanylyltransferase
GTase
mRNA guanylyltransferase
EC 2.7.7.50
PF03919 (mRNA_cap_C)
PF01331 (mRNA_cap_enzyme)
[Graphical View]

KEGG enzyme name
mRNA guanylyltransferase
mRNA capping enzyme
messenger RNA guanylyltransferase
Protein 2

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
Q84424 MCE_PBCV1 GTP + (5'')pp-Pur-mRNA = diphosphate + G(5'')ppp-Pur-mRNA. Monomer. Magnesium or manganese.
P78587 MCE1_CANAL GTP + (5'')pp-Pur-mRNA = diphosphate + G(5'')ppp-Pur-mRNA. The mRNA-capping enzyme is composed of two separate chains alpha and beta, respectively a mRNA guanylyltransferase and an RNA 5''-triphosphatase. Nucleus.

KEGG Pathways
Map code Pathways E.C.

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00305 C00034 C00044 C02100 C00013 C02031
E.C.
Compound Magnesium Manganese GTP (5')ppPur-mRNA Pyrophosphate G(5')pppR-RNA GMP covalently bonded to Lys82
Type divalent metal (Ca2+, Mg2+) heavy metal amide group,amine group,nucleotide nucleic acids,phosphate group/phosphate ion phosphate group/phosphate ion amide group,amine group,nucleic acids,nucleotide
ChEBI 18420
 18420
 		18291
 35154
 18291
 35154
 		15996
 15996
 		29888
 29888
PubChem 888
 888
 		23930
 23930
 		6830
 6830
 		1023
 21961011
 1023
 21961011
1ckmA01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ckmB01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1cknA01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1cknB01 Unbound Bound:_MN Unbound Unbound Analogue:SO4 Unbound Unbound
1ckoA01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1p16A01 Unbound Unbound Unbound Unbound Analogue:PO4_7001 Unbound Unbound
1p16B01 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ckmA02 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ckmB02 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1cknA02 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1cknB02 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ckoA02 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1p16A03 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1p16B03 Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ckmA03 Unbound Unbound Bound:GTP Unbound Unbound Unbound Unbound
1ckmB03 Unbound Unbound Bound:GTP Unbound Unbound Unbound Unbound
1cknA03 Unbound Unbound Bound:GTP Unbound Unbound Unbound Unbound
1cknB03 Unbound Unbound Unbound Unbound Unbound Unbound Bound:GPL
1ckoA03 Unbound Unbound Unbound Unbound Unbound Bound:GP3 Unbound
1p16A02 Unbound Unbound Unbound Unbound Unbound Unbound Bound:__G
1p16B02 Unbound Unbound Bound:GTP Unbound Unbound Unbound Unbound

Reference for Active-site residues
resource references E.C.
literature [11]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1ckmA01 ARG 106
1ckmB01 ARG 106
1cknA01 ARG 106
1cknB01 ARG 106
1ckoA01 ARG 106
1p16A01 ARG 92
1p16B01 ARG 92
1ckmA02 LYS 236;ASP 244;ARG 295;LYS 298;ASN 302
1ckmB02 LYS 236;ASP 244;ARG 295;LYS 298;ASN 302
1cknA02 LYS 236;ASP 244;ARG 295;LYS 298;ASN 302
1cknB02 LYS 236;ASP 244;ARG 295;LYS 298;ASN 302
1ckoA02 LYS 236;ASP 244;ARG 295;LYS 298;ASN 302
1p16A03 LYS 243;ASP 251;ARG 342;LYS 345;ASN 349
1p16B03 LYS 243;ASP 251;ARG 342;LYS 345;ASN 349
1ckmA03 LYS 82;ASP 213;LYS 234
1ckmB03 LYS 82;ASP 213;LYS 234
1cknA03 LYS 82;ASP 213;LYS 234
1cknB03 ;ASP 213;LYS 234 GPL 82 GPL guanylated lysine
1ckoA03 LYS 82;ASP 213;LYS 234
1p16A02 LYS 67;ASP 220;LYS 241
1p16B02 LYS 67;ASP 220;LYS 241

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[8]
Fig.7, p.549-552
[10]
p.9575-9576
[11]
Fig.4, p.1508-1509
[15]
[16]
p.1556-1558
[17]
p.761-762

References
[1]
Resource
Comments
Medline ID
PubMed ID 2159008
Journal J Biol Chem
Year 1990
Volume 265
Pages 7669-72
Authors Fausnaugh J, Shatkin AJ
Title Active site localization in a viral mRNA capping enzyme.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID 2161527
Journal Proc Natl Acad Sci U S A
Year 1990
Volume 87
Pages 4023-7
Authors Guo PX, Moss B
Title Interaction and mutual stabilization of the two subunits of vaccinia virus mRNA capping enzyme coexpressed in Escherichia coli.
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID 8385101
Journal J Biol Chem
Year 1993
Volume 268
Pages 7256-60
Authors Cong P, Shuman S
Title Covalent catalysis in nucleotidyl transfer. A KTDG motif essential for enzyme-GMP complex formation by mRNA capping enzyme is conserved at the active sites of RNA and DNA ligases.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 8227060
Journal J Biol Chem
Year 1993
Volume 268
Pages 24986-9
Authors Niles EG, Christen L
Title Identification of the vaccinia virus mRNA guanyltransferase active site lysine.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 8195132
Journal J Biol Chem
Year 1994
Volume 269
Pages 14974-81
Authors Higman MA, Christen LA, Niles EG
Title The mRNA (guanine-7-)methyltransferase domain of the vaccinia virus mRNA capping enzyme. Expression in Escherichia coli and structural and kinetic comparison to the intact capping enzyme.
Related PDB
Related UniProtKB
[6]
Resource
Comments
Medline ID
PubMed ID 7991582
Journal Proc Natl Acad Sci U S A
Year 1994
Volume 91
Pages 12046-50
Authors Shuman S, Liu Y, Schwer B
Title Covalent catalysis in nucleotidyl transfer reactions: essential motifs in Saccharomyces cerevisiae RNA capping enzyme are conserved in Schizosaccharomyces pombe and viral capping enzymes and among polynucleotide ligases.
Related PDB
Related UniProtKB
[7]
Resource
Comments
Medline ID
PubMed ID 7565775
Journal Mol Cell Biol
Year 1995
Volume 15
Pages 6222-31
Authors Cong P, Shuman S
Title Mutational analysis of mRNA capping enzyme identifies amino acids involved in GTP binding, enzyme-guanylate formation, and GMP transfer to RNA.
Related PDB
Related UniProtKB
[8]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.5 ANGSTROMS)
Medline ID 97304383
PubMed ID 9160746
Journal Cell
Year 1997
Volume 89
Pages 545-53
Authors Hakansson K, Doherty AJ, Shuman S, Wigley DB
Title X-ray crystallography reveals a large conformational change during guanyl transfer by mRNA capping enzymes.
Related PDB 1ckm 1ckn
Related UniProtKB Q84424
[9]
Resource
Comments
Medline ID
PubMed ID 9371657
Journal J Virol
Year 1997
Volume 71
Pages 9837-43
Authors Yu L, Martins A, Deng L, Shuman S
Title Structure-function analysis of the triphosphatase component of vaccinia virus mRNA capping enzyme.
Related PDB
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID 9275164
Journal Proc Natl Acad Sci U S A
Year 1997
Volume 94
Pages 9573-8
Authors Wang SP, Deng L, Ho CK, Shuman S
Title Phylogeny of mRNA capping enzymes.
Related PDB
Related UniProtKB
[11]
Resource
Comments X-RAY CRYSTALLOGRAPHY (3.1 ANGSTROMS) OF 11-327
Medline ID 98132620
PubMed ID 9465045
Journal Proc Natl Acad Sci U S A
Year 1998
Volume 95
Pages 1505-10
Authors Hakansson K, Wigley DB
Title Structure of a complex between a cap analogue and mRNA guanylyl transferase demonstrates the structural chemistry of RNA capping.
Related PDB 1cko
Related UniProtKB Q84424
[12]
Resource
Comments
Medline ID
PubMed ID 10454631
Journal Nucleic Acids Res
Year 1999
Volume 27
Pages 3253-8
Authors Doherty AJ
Title Conversion of a DNA ligase into an RNA capping enzyme.
Related PDB
Related UniProtKB
[13]
Resource
Comments
Medline ID
PubMed ID 11018011
Journal Genes Dev
Year 2000
Volume 14
Pages 2435-40
Authors Schroeder SC, Schwer B, Shuman S, Bentley D
Title Dynamic association of capping enzymes with transcribing RNA polymerase II.
Related PDB
Related UniProtKB
[14]
Resource
Comments
Medline ID
PubMed ID 11463793
Journal J Biol Chem
Year 2001
Volume 276
Pages 36116-24
Authors Hausmann S, Ho CK, Schwer B, Shuman S
Title An essential function of Saccharomyces cerevisiae RNA triphosphatase Cet1 is to stabilize RNA guanylyltransferase Ceg1 against thermal inactivation.
Related PDB
Related UniProtKB
[15]
Resource
Comments
Medline ID
PubMed ID 12846573
Journal Biochemistry
Year 2003
Volume 42
Pages 8240-9
Authors Sawaya R, Shuman S
Title Mutational analysis of the guanylyltransferase component of Mammalian mRNA capping enzyme.
Related PDB
Related UniProtKB
[16]
Resource
Comments
Medline ID
PubMed ID 12820968
Journal Mol Cell
Year 2003
Volume 11
Pages 1549-61
Authors Fabrega C, Shen V, Shuman S, Lima CD
Title Structure of an mRNA capping enzyme bound to the phosphorylated carboxy-terminal domain of RNA polymerase II.
Related PDB
Related UniProtKB
[17]
Resource
Comments
Medline ID
PubMed ID 15582400
Journal Curr Opin Struct Biol
Year 2004
Volume 14
Pages 757-64
Authors Shuman S, Lima CD
Title The polynucleotide ligase and RNA capping enzyme superfamily of covalent nucleotidyltransferases.
Related PDB
Related UniProtKB

Comments
According to the literature [8], [10], [11], the catalytic reaction probably proceeds as follows:
(1) Asp213 acts as a general base, which deprotonates the nucleophile, the sidechain of Lys82, to activate it. (Although the literature [11] suggests an alternative mechanism, in which alpha-phosphate of GTP may act as a general base to deprotonate Lys82 and Asp213 may only modulate it by hydrogen bonding, there some factors against the substrate-assisted mechanism. Cofactor magnesium/manganese ion might neutralize the negative charge changing the pKa of the phosphate group. Moreover, the proton transfer from Lys82 to the phosphate results in the loss of an ion pair, which will not be favored. see [11])
(2) The sidechain of Lys82 makes a nucleophilic attack on the alpha-phosphate group of GTP. This process leads to the transition state in which the transferred phosphate group goes through a pentacovalent state (SN2-like reaction). (During this process, the domains are in the closed conformation.)
(3) The transferred alpha-phosphate group is stabilized by Lys234 and Lys236 along with a cofactor magnesium/manganese ion. Meanwhile, Arg106, Arg295, Lys298 and Asn302 stabilize the leaving beta- and gamma-phosphate groups, along with Asp244, which might neutralize these positively charged residues.
(4) Phosphoamide intermediate is formed between Lys82 and GMP, whereas the leaving beta- and gamma-phosphate is released from the enzyme as a product, diphosphate.
(5) The terminal phosphate group of the second substrate, (5')ppPur-mRNA, is bound to the active site. (During this process, the enzyme is in the open conformation.)
(6) The oxygen atom of the terminal phosphate group makes a nucleophilic attack on the phosphorus atom of GMP attached to Lys82. This process leads to the transition state in which the transferred phosphate group goes through a pentacovalent state (SN2-like reaction).
(7) The transferred phosphate group is again stabilized by Lys234 and Lys236 along with a cofactor magnesium/manganese ion. Meanwhile, Arg106, Arg295, Lys298 and Asn302 stabilize the acceptor phosphate groups, along with Asp244, which might neutralize these positively charged residues.
(8) Asp213 acts as a general acid to protonate Lys82, which is finally released from the product.

Created Updated
2004-03-25 2009-02-26