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PDBsum entry 3hdf
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References listed in PDB file
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Key reference
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Title
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Regulation of a muralytic enzyme by dynamic membrane topology.
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Authors
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Q.Sun,
G.F.Kuty,
A.Arockiasamy,
M.Xu,
R.Young,
J.C.Sacchettini.
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Ref.
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Nat Struct Biol, 2009,
16,
1192-1194.
[DOI no: ]
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PubMed id
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Abstract
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R(21), the lysozyme of coliphage 21, has an N-terminal signal-anchor-release
(SAR) domain that directs its secretion in a membrane-tethered, inactive form
and then its release and activation in the periplasm. Both genetic and
crystallographic studies show that the SAR domain, once extracted from the
bilayer, refolds into the body of the enzyme and effects muralytic activation by
repositioning one residue of the canonical lysozyme catalytic triad.
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Figure 1.
(a) N-terminal sequences of R^21, Lyz^P1 and T4 E. The
N-terminal domains of R^21, Lyz^P1 and T4 E are shown aligned by
their Glu-8aa-Asp/Cys-5aa-Thr catalytic triad (blue and
asterisks). SAR domains are boxed in orange. Leucine
substitutions made in R^21 are shown above Gly14 and Gly15. The
PelB signal sequence (PelB[ss]) and the artificial transmembrane
domain (TMD[art]; yellow) are shown with arrows indicating
points of fusion in the chimeric constructs. Residues involved
in positioning the catalytic glutamate are highlighted by a
light blue box in Lyz^P1 and T4 E. (b) Lysis profiles. (  )
pZE-luc, (  circle
) pZE-R^21[G14,15L], (  )
pZE-luc + 1 mM CHCl[3] at 80 min after induction, (  square
) pZE-R^21[G14,15L] + 1 mM CHCl[3] at 80 min after induction, (
 )
pZE-R^21, (  diamond
)Lyz^P1[1–26]  R^21[27–165],
(  )
pZE-pelB[ss] R^21[27–165],
(  triangle
) pZE-TMD[art] R^21.
A[550], absorbance at 550 nm. (c) Localization and processing of
R^21 and derivatives after expression of the indicated chimeras.
In top panel, lanes 1, 2 and 3 represent the total (T),
periplasm (P) and spheroplast (S) fractions, respectively.
Below, lanes 1, 2 and 3 represent the total (T), soluble (S) and
membrane (M) fractions, respectively. (d) Morphologies of cells
expressing the indicated pZE-R^21 plasmids at 100 min after
induction. Scale bars (throughout) are 5  m.
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Figure 2.
(a) Topological and conformational dynamics of R^21
activation. Crystal structures of ^iR^21 (left) and ^aR^21
(right) are represented in cartoon format, with  -helix
in cyan,  -strand
in magenta and coil in salmon; the SAR domain is shown in orange
and is depicted as a membrane-spanning helix in ^iR^21. The
catalytic triads are represented in stick-and-ball form, with
disulfide linkages in yellow stick. (b) Alignment of the
catalytic loop regions of ^iR^21 (green) and ^aR^21 (salmon).
Except for Glu35, which shows a 10-Å C displacement
(dashed line), most of the catalytic loop (Ser38–Thr67) of
^aR^21 can be superimposed on the same region of ^iR^21 (r.m.s.
difference = 0.4 Å for 209 atoms). (c) Alignment of the
C-terminal domains of ^iR^21 and ^aR^21, colored as in b.
Beginning at Glu96, the backbone r.m.s. difference between the
two structures is 4.0 Å. Helix 8
in ^iR^21 tilts 30°, turns and unwinds compared to ^aR^21,
thereby turning Arg152 away from Glu35. (d) Polarity switching
at the interface for the SAR domain. The calculated
electrostatic surface (positive = blue; negative = red)
contacting the helices of the extracted SAR domain for ^aR^21 is
shown at right; the corresponding surface is ^iR^21 is shown at
left, with the SAR helices superimposed as an orange backbone
ribbon trace.
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nat Struct Biol
(2009,
16,
1192-1194)
copyright 2009.
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