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CSA entry for 1fa0
Original Entry
Title:
Transferase
Compound:
Poly(a)-polymerase
Mutant:
No
UniProt/Swiss-Prot:
P29468-PAP_YEAST
EC Class:
2.7.7.19
Other CSA Entries:
Overview of all sites for 1fa0
Homologues of 1fa0
Entries for UniProt/Swiss-Prot: P29468
Entries for EC: 2.7.7.19
Other Databases:
PDB entry: 1fa0
PDBsum entry: 1fa0
UniProt/Swiss-Prot: P29468
IntEnz entry: 2.7.7.19
Literature Report:
Introduction:
Poly(A) polymerase (PAP) catalyses the addition of a poly-adenosine tail to almost all eukaryotic mRNAs. This poly(A) tail has numerous functions in eukaryotes: it facilitates transport of the mRNA from the nucleus, regulates mRNA stability, and increases the efficiency of translation. Poly(A) tails are added to mRNA by a multiprotein complex that recognises the polyadenylation signal, cleaves the precursor mRNA, and adds the additional nucleotides. The poly(A) polymerase component retains its polymerase activity when isolated from the holoenzyme assembly and will processively add long stretches of adenosine nucleotides to an RNA primer in vitro.
Mechanism:
Poly(A) polymerase is thought to use the two-metal-ion mechanism proposed for all nucleic acid polymerases. One metal ion, dubbed Metal A (MN 601 in structure 1fa0) contacts the 3' OH of the primer and lowers its pKa, activating it for in-line nucleophilic attack on the alpha phosphate of the incoming ATP. Accumulation of negative charge on the alpha phosphate in the pentacoordinate transition state is stabilised by both Metal A and Metal B (MN 600 in structure 1fa0). Metal B additionally functions to stabilise accumulation of negative charge on the pyrophosphate leaving group. Lys 215 may also stabilise negative charge on the pyrophosphate leaving group.
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Found by:
Literature reference 

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
LYSA 215 215Sidechain
ElectrostaticTransition state
Proposed to stabilise the negatively charged pyrophosphate leaving group.
Evidence from paper Evidence concerns Evidence type
PubMed ID 10958780 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 10958780 Current protein Structural similarity to homologue of known mechanism

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
MNA 600 0
ElectrostaticTransition state
Stabilises negative charge on the alpha phosphate in the pentacoordinate transition state. Stabilises accumulation of negative charge on the pyrophosphate leaving group.
Evidence from paper Evidence concerns Evidence type
PubMed ID 10958780 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 10958780 Current protein Structural similarity to homologue of known mechanism

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
MNA 601 0
ElectrostaticTransition state
ElectrostaticSubstrate
Coordinates the 3' OH of the primer and lowers its pKa, promoting formation of the nucleophilic oxyanion. Coordinates the alpha phosphate of the ATP and stabilises negative charge in the pentacoordinate transition state.
Evidence from paper Evidence concerns Evidence type
PubMed ID 10958780 Current protein Structural similarity to homologue of known mechanism
PubMed ID 10958780 Current protein Residue is positioned appropriately (ligand position known)
References:
1
Structure of yeast poly(A) polymerase alone and in complex with 3'-dATP.
J. Bard and A. M. Zhelkovsky and S. Helmling and T. N. Earnest and C. L. Moore and A. Bohm
Science 289, (5483) 1346-9, (2000).
10958780
2
Structural and functional insights provided by crystal structures of DNA polymerases and their substrate complexes.
C. A. Brautigam and T. A. Steitz
Curr Opin Struct Biol 8, (1) 54-63, (1998).
9519297
3
The mechanism of action of T7 DNA polymerase.
S. DoubliƩ and T. Ellenberger
Curr Opin Struct Biol 8, (6) 704-12, (1998).
9914251
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