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PDBsum entry 3fy3
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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 and functional studies of truncated hemolysin a from proteus mirabilis.
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Authors
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T.M.Weaver,
J.M.Hocking,
L.J.Bailey,
G.T.Wawrzyn,
D.R.Howard,
L.A.Sikkink,
M.Ramirez-Alvarado,
J.R.Thompson.
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Ref.
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J Biol Chem, 2009,
284,
22297-22309.
[DOI no: ]
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PubMed id
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Note: In the PDB file this reference is
annotated as "TO BE PUBLISHED". The citation details given above have
been manually determined.
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Abstract
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In this study we analyzed the structure and function of a truncated form of
hemolysin A (HpmA265) from Proteus mirabilis using a series of functional and
structural studies. Hemolysin A belongs to the two-partner secretion pathway.
The two-partner secretion pathway has been identified as the most common protein
secretion pathway among Gram-negative bacteria. Currently, the mechanism of
action for the two-partner hemolysin members is not fully understood. In this
study, hemolysis experiments revealed a unidirectional, cooperative, biphasic
activity profile after full-length, inactive hemolysin A was seeded with
truncated hemolysin A. We also solved the first x-ray structure of a TpsA
hemolysin. The truncated hemolysin A formed a right-handed parallel beta-helix
with three adjoining segments of anti-parallel beta-sheet. A CXXC disulfide
bond, four buried solvent molecules, and a carboxyamide ladder were all located
at the third complete beta-helix coil. Replacement of the CXXC motif led to
decreased activity and stability according to hemolysis and CD studies.
Furthermore, the crystal structure revealed a sterically compatible, dry dimeric
interface formed via anti-parallel beta-sheet interactions between neighboring
beta-helix monomers. Laser scanning confocal microscopy further supported the
unidirectional interconversion of full-length hemolysin A. From these results, a
model has been proposed, where cooperative, beta-strand interactions between
HpmA265 and neighboring full-length hemolysin A molecules, facilitated in part
by the highly conserved CXXC pattern, account for the template-assisted
hemolysis.
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Figure 2.
The crystal structures of HpmA265, Fha30, and HMW1-PP show
differences within the top and flanking anti-parallel β-sheet
regions.a, the Cα coordinates have been superimposed between
HpmA265 and Fha30. The trace for HpmA Cα atoms has been colored
black in both images. Structural differences between HpmA265 and
Fha30 have been colored cyan for the top and flanking regions
apβ1, apβ2, and apβ3, while differences within β-arc regions
have been colored red. The largest structural differences
between HpmA265 and Fha30 lie at the flanking anti-parallel
sheet apβ3. This shift accommodates a 12-residue insertion
between β22 and β23 in HpmA265 and preserves the integrity of
the β-helix core alignment. b, structural differences between
HpmA265 and HMW1-PP have been colored cyan for anti-parallel
regions, magenta for α-helix regions, and red for β-arcs. The
large shift at the N-terminal end derives from a global
superposition of three anti-parallel strands within HMW1-PP onto
the four anti-parallel strands within HpmA265. This large
difference stems from moving β2 of HMW1-PP between β1 and β2
of HpmA265. The N-terminal superposition also leaves β5 and β6
of HpmA265 without matching HMW1-PP strands. Additionally,
HMW1-PP has an α-helix in place of two of the anti-parallel
strands within apβ3. This creates another location of large
structural difference. In both instances, the structural
agreement between β-helix core Cα atoms is well maintained,
especially within the NPNG motif (colored green).
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Figure 9.
HpmA265 crystallographic dry dimer interface leading to a
filamentous appearance.a, the HpmA265 dry dimer forms a
filamentous structure. The solid lines represent hydrogen bonds
shared between β23 strands of both subunits. b, the dry dimer
interface has been displayed where β23 from each monomer has
been colored independently. The anti-parallel interstrand
hydrogen-bonding network creates dry, sterically compatible
dimer interface. Dashed lines represent hydrogen bonds, while
van der Waal's surfaces have been provided for various
hydrophobic side chains.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2009,
284,
22297-22309)
copyright 2009.
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