 |
PDBsum entry 3d6y
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Transcription regulator/DNA
|
PDB id
|
|
|
|
3d6y
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
Structures of bmrr-Drug complexes reveal a rigid multidrug binding pocket and transcription activation through tyrosine expulsion.
|
|
|
Authors
|
|
K.J.Newberry,
J.L.Huffman,
M.C.Miller,
N.Vazquez-Laslop,
A.A.Neyfakh,
R.G.Brennan.
|
|
|
Ref.
|
|
J Biol Chem, 2008,
283,
26795-26804.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
BmrR is a member of the MerR family and a multidrug binding transcription factor
that up-regulates the expression of the bmr multidrug efflux transporter gene in
response to myriad lipophilic cationic compounds. The structural mechanism by
which BmrR binds these chemically and structurally different drugs and
subsequently activates transcription is poorly understood. Here, we describe the
crystal structures of BmrR bound to rhodamine 6G (R6G) or berberine (Ber) and
cognate DNA. These structures reveal each drug stacks against multiple aromatic
residues with their positive charges most proximal to the carboxylate group of
Glu-253 and that, unlike other multidrug binding pockets, that of BmrR is rigid.
Substitution of Glu-253 with either alanine (E253A) or glutamine (E253Q) results
in unpredictable binding affinities for R6G, Ber, and tetraphenylphosphonium.
Moreover, these drug binding studies reveal that the negative charge of Glu-253
is not important for high affinity binding to Ber and tetraphenylphosphonium but
plays a more significant, but unpredictable, role in R6G binding. In vitro
transcription data show that E253A and E253Q are constitutively active, and
structures of the drug-free E253A-DNA and E253Q-DNA complexes support a
transcription activation mechanism requiring the expulsion of Tyr-152 from the
multidrug binding pocket. In sum, these data delineate the mechanism by which
BmrR binds lipophilic, monovalent cationic compounds and suggest the importance
of the redundant negative electrostatic nature of this rigid drug binding pocket
that can be used to discriminate against molecules that are not substrates of
the Bmr multidrug efflux pump.
|
|
|
|
|
|
|
|
Figure 2.
FIGURE 2. Drug binding to BmrR. Key residues are labeled
and shown as pale blue sticks with the oxygen atoms colored red
and nitrogen atoms colored blue. Secondary structural elements
are depicted as pale blue ribbons (coils,  helices; arrows, β
strands). A, wtBmrR-imidazole-DNA complex (21). Imidazole
molecules are shown as green sticks, and dashed lines denote
hydrogens bonds. B, wtBmrR-R6G-DNA complex. R6G is shown as red
sticks with its positive charge depicted by a red +.
Electrostatic interactions are shown as dashed lines. C,
wtBmrR-Ber-DNA complex. Ber is shown as yellow sticks, and its
positive charge depicted by a blue +. Electrostatic interactions
are shown as dashed lines. D, superposition of the
BmrR-imidazole-DNA and BmrR-R6G-DNA complexes. E, superposition
of all BmrR-drug-DNA complexes. TPP^+ is shown as green sticks
with its positively charged phosphorus atom colored purple. F,
superposition of E253Q-DNA and BmrR-R6G-DNA structures.
E253Q-DNA is shown as green ribbons and sticks. Dashed lines
indicate potential clashes between R6G and residues in the E253Q
structure. Figs. 2, 3, 4, 5 and 6 were made with PyMol (Delano,
W. L. (2002)).
|
|
Figure 6.
FIGURE 6. Electrostatic surface representation of wild type
and mutant BmrR drug binding pockets. Electropositive surfaces
are shown in blue, and electronegative surfaces are shown in
red. The left panel shows the electrostatic surface potential of
the wtBmrR-R6G-DNA complex with R6G depicted as cyan sticks
within the pocket. The middle panel depicts the E253A surface,
and the right panel the E253Q surface.
|
|
|
|
|
|
The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2008,
283,
26795-26804)
copyright 2008.
|
|
|
|
|
|
|
