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PDBsum entry 3d4b
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References listed in PDB file
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Key reference
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Title
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Structural insights into intermediate steps in the sir2 deacetylation reaction.
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Authors
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W.F.Hawse,
K.G.Hoff,
D.G.Fatkins,
A.Daines,
O.V.Zubkova,
V.L.Schramm,
W.Zheng,
C.Wolberger.
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Ref.
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Structure, 2008,
16,
1368-1377.
[DOI no: ]
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PubMed id
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Abstract
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Sirtuin enzymes comprise a unique class of NAD(+)-dependent protein
deacetylases. Although structures of many sirtuin complexes have been
determined, structural resolution of intermediate chemical steps are needed to
understand the deacetylation mechanism. We report crystal structures of the
bacterial sirtuin, Sir2Tm, in complex with an S-alkylamidate intermediate,
analogous to the naturally occurring O-alkylamidate intermediate, and a Sir2Tm
ternary complex containing a dissociated NAD(+) analog and acetylated peptide.
The structures and biochemical studies reveal critical roles for the invariant
active site histidine in positioning the reaction intermediate, and for a
conserved phenylalanine residue in shielding reaction intermediates from base
exchange with nicotinamide. The new structural and biochemical studies provide
key mechanistic insight into intermediate steps of the Sir2 deacetylation
reaction.
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Figure 1.
Figure 1. Sirtuin Deacetylation Reaction Mechanism
(A
and B) (A) In the first step of the Sir2 deacetylation reaction
(I), the ADP-ribose moiety of NAD^+ is transferred to
acetyl-lysine, generating the O-alkylamidate intermediate. In
this step (I), the nicotinamide-N-ribose bond is broken to
generate free nicotinamide. Next, the N-ribose 2′OH group
attacks the O-alkylamidate intermediate, generating a 1′,2′
bicyclic species (II). Subsequent hydrolysis of the 1′,2′
bicyclic species yields deacetylated lysine and
2′O-acetyl-ADP-ribose (III). The structure of the DADMe-NAD^+
analog, which represents a dissociated NAD^+ species, is
depicted in (B).
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Figure 5.
Figure 5. A Conserved Cluster of Phenylalanines Protects the
Peptidyl-Imidate Intermediate from Hydrolysis and Base Exchange
with Nicotinamide
(A) Cluster of phenylalanine side chains
(red), Phe33, Phe48, and Phe162, that shield the S-alkylamidate
intermediate from solvent. (B) Orientation of Phe33 side chain
in the Michaelis complex (blue) and in the structure containing
the S-alkylamidate intermediate (grey). (C) Conformation of Phe
33 in the Sir2Tm-S-alkylamidate intermediate (pink) as compared
with its position in the structure of Sir2Tm bound to the
deacetylation reaction product, nicotinamid (green; PDB ID code
2H4J).
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The above figures are
reprinted
from an Open Access publication published by Cell Press:
Structure
(2008,
16,
1368-1377)
copyright 2008.
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