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PDBsum entry 4k4c
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References listed in PDB file
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Key reference
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Title
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Structure and catalysis in the escherichia coli hotdog-Fold thioesterase paralogs ydii and ybdb.
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Authors
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R.Wu,
J.A.Latham,
D.Chen,
J.Farelli,
H.Zhao,
K.Matthews,
K.N.Allen,
D.Dunaway-Mariano.
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Ref.
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Biochemistry, 2014,
53,
4788-4805.
[DOI no: ]
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PubMed id
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Abstract
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Herein, the structural determinants for substrate recognition and catalysis in
two hotdog-fold thioesterase paralogs, YbdB and YdiI from Escherichia coli, are
identified and analyzed to provide insight into the evolution of biological
function in the hotdog-fold enzyme superfamily. The X-ray crystal structures of
YbdB and YdiI, in complex with inert substrate analogs, determined in this study
revealed the locations of the respective thioester substrate binding sites and
the identity of the residues positioned for substrate binding and catalysis. The
importance of each of these residues was assessed through amino acid
replacements followed by steady-state kinetic analyses of the corresponding
site-directed mutants. Transient kinetic and solvent (18)O-labeling studies were
then carried out to provide insight into the role of Glu63 posited to function
as the nucleophile or general base in catalysis. Finally, the
structure-function-mechanism profiles of the two paralogs, along with that of a
more distant homolog, were compared to identify conserved elements of substrate
recognition and catalysis, which define the core traits of the hotdog-fold
thioesterase family, as well as structural features that are unique to each
thioesterase. Founded on the insight gained from this analysis, we conclude that
the promiscuity revealed by in vitro substrate activity determinations, and
posited to facilitate the evolution of new biological function, is the product
of intrinsic plasticity in substrate binding as well as in the catalytic
mechanism.
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