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PDBsum entry 2bf5
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Oxidoreductase
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PDB id
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2bf5
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References listed in PDB file
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Key reference
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Title
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Crystal structures and functional studies of t4mod, The toluene 4-Monooxygenase catalytic effector protein.
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Authors
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G.T.Lountos,
K.H.Mitchell,
J.M.Studts,
B.G.Fox,
A.M.Orville.
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Ref.
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Biochemistry, 2005,
44,
7131-7142.
[DOI no: ]
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PubMed id
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Abstract
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Toluene 4-monooxygenase (T4MO) is a four-component complex that catalyzes the
regiospecific, NADH-dependent hydroxylation of toluene to yield p-cresol. The
catalytic effector (T4moD) of this complex is a 102-residue protein devoid of
metals or organic cofactors. It forms a complex with the diiron hydroxylase
component (T4moH) that influences both the kinetics and regiospecificity of
catalysis. Here, we report crystal structures for native T4moD and two
engineered variants with either four (DeltaN4-) or 10 (DeltaN10-) residues
removed from the N-terminal at 2.1-, 1.7-, and 1.9-A resolution, respectively.
The crystal structures have C-alpha root-mean-squared differences of less than
0.8 A for the central core consisting of residues 11-98, showing that
alterations of the N-terminal have little influence on the folded core of the
protein. The central core has the same fold topology as observed in the NMR
structures of T4moD, the methane monooxygenase effector protein (MmoB) from two
methanotrophs, and the phenol hydroxylase effector protein (DmpM). However, the
root-mean-squared differences between comparable C-alpha positions in the X-ray
structures and the NMR structures vary from approximately 1.8 A to greater than
6 A. The X-ray structures exhibit an estimated overall coordinate error from
0.095 (0.094) A based on the R-value (R free) for the highest resolution
DeltaN4-T4moD structure to 0.211 (0.196) A for the native T4moD structure.
Catalytic studies of the DeltaN4-, DeltaN7-, and DeltaN10- variants of T4moD
show statistically insignificant changes in k(cat), K(M), k(cat)/K(M), and K(I)
relative to the native protein. Moreover, there was no significant change in the
regiospecificity of toluene oxidation with any of the T4moD variants. The
relative insensitivity to changes in the N-terminal region distinguishes T4moD
from the MmoB homologues, which each require the approximately 33 residue
N-terminal region for catalytic activity.
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Secondary reference #1
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Title
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Crystallization and preliminary analysis of native and n-Terminal truncated isoforms of toluene-4-Monooxygenase catalytic effector protein.
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Authors
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A.M.Orville,
J.M.Studts,
G.T.Lountos,
K.H.Mitchell,
B.G.Fox.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2003,
59,
572-575.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 Toluene 4-monooxygenase enzyme complex. The catalytic
effector protein (T4moD, 11.6 kDa) is the subject of this work.
The natural enzyme complex produces >95% yield of p-cresol from
NADH, O[2] and toluene (Mitchell et al., 2002[Mitchell, K. H.,
Studts, J. M. & Fox, B. G. (2002). Biochemistry, 41,
3176-3188.]).
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The above figure is
reproduced from the cited reference
with permission from the IUCr
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Secondary reference #2
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Title
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Solution structure of the toluene 4-Monooxygenase effector protein (t4mod).
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Authors
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H.Hemmi,
J.M.Studts,
Y.K.Chae,
J.Song,
J.L.Markley,
B.G.Fox.
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Ref.
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Biochemistry, 2001,
40,
3512-3524.
[DOI no: ]
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PubMed id
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Secondary reference #3
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Title
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Application of fed-Batch fermentation to the preparation of isotopically labeled or selenomethionyl-Labeled proteins.
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Authors
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J.M.Studts,
B.G.Fox.
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Ref.
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Protein Expr Purif, 1999,
16,
109-119.
[DOI no: ]
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PubMed id
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Secondary reference #4
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Title
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Combined participation of hydroxylase active site residues and effector protein binding in a para to ortho modulation of toluene 4-Monooxygenase regiospecificity.
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Authors
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K.H.Mitchell,
J.M.Studts,
B.G.Fox.
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Ref.
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Biochemistry, 2002,
41,
3176-3188.
[DOI no: ]
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PubMed id
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Secondary reference #5
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Title
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Recombinant toluene-4-Monooxygenase: catalytic and mössbauer studies of the purified diiron and rieske components of a four-Protein complex.
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Authors
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J.D.Pikus,
J.M.Studts,
C.Achim,
K.E.Kauffmann,
E.Münck,
R.J.Steffan,
K.Mcclay,
B.G.Fox.
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Ref.
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Biochemistry, 1996,
35,
9106-9119.
[DOI no: ]
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PubMed id
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