 |
PDBsum entry 2p5t
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Transcription regulator
|
PDB id
|
|
|
|
2p5t
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
Molecular and structural characterization of the pezat chromosomal toxin-Antitoxin system of the human pathogen streptococcus pneumoniae.
|
|
|
Authors
|
|
S.K.Khoo,
B.Loll,
W.T.Chan,
R.L.Shoeman,
L.Ngoo,
C.C.Yeo,
A.Meinhart.
|
|
|
Ref.
|
|
J Biol Chem, 2007,
282,
19606-19618.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
The chromosomal pezT gene of the Gram-positive pathogen Streptococcus pneumoniae
encodes a protein that is homologous to the zeta toxin of the Streptococcus
pyogenes plasmid pSM19035-encoded epsilon-zeta toxin-antitoxin system.
Overexpression of pezT in Escherichia coli led to severe growth inhibition from
which the bacteria recovered approximately 3 h after induction of expression.
The toxicity of PezT was counteracted by PezA, which is encoded immediately
upstream of pezT and shares weak sequence similarities in the C-terminal region
with the epsilon antitoxin. The pezAT genes form a bicistronic operon that is
co-transcribed from a sigma(70)-like promoter upstream of pezA and is negatively
autoregulated with PezA functioning as a transcriptional repressor and PezT as a
co-repressor. Both PezA and the non-toxic PezA(2)PezT(2) protein complex bind to
a palindrome sequence that overlaps the promoter. This differs from the
epsilon-zeta system in which epsilon functions solely as the antitoxin and
transcriptional regulation is carried out by another protein designated omega.
Results from site-directed mutagenesis experiments demonstrated that the
toxicity of PezT is dependent on a highly conserved phosphoryltransferase active
site and an ATP/GTP nucleotide binding site. In the PezA(2)PezT(2) complex, PezA
neutralizes the toxicity of PezT by blocking the nucleotide binding site through
steric hindrance.
|
|
|
|
|
|
|
|
Figure 1.
FIGURE 1. A, sequence alignment of the C-terminal domain of
PezA and PezT with the epsilon/zeta system. Identical residues
are colored green, highly homologous residues orange, and
homologous residues yellow, respectively. Secondary structures
are shown above the sequence alignment with  -helices illustrated as
cylinders and  -strands as arrows. The
region in PezT that could not be modeled into the electron
density is indicated by a green line. Residues that are involved
in PezA-PezA interaction are labeled with filled squares beneath
the sequence alignment, and those that are involved in PezA-PezT
interaction with filled triangles. Open circles label residues
of PezA that inhibit the toxin PezT. Asterisks indicate those
residues that were changed by site-directed mutagenesis in PezT
and were shown to abolish PezT toxicity. Filled diamonds above
the sequence alignment indicate residues of different PezT
variants where mutations to homologous residues were observed,
open diamonds mark mutations of a non-conservative nature. B,
ribbon representation of the heterotetrameric PezA/PezT
arrangement. Helices of PezA are colored in blue and helices of
PezT in beige, respectively. -Strands in PezT are
colored in red. C, surface representation of the complex colored
according to the color scheme of A.
|
|
Figure 6.
FIGURE 6. A, surface representation of PezT alone, colored
according to Fig. 1A. Residues that are identical with zeta and
showed to impair PezT toxicity are colored in blue. An ATP
molecule colored in magenta was modeled according to the
position found in the chloramphenicol-phosphotransferase (29).
B, effect of PezT mutations on the growth of E. coli
BL21(DE3)/pLysS carrying recombinant pET11a plasmids as
determined by A[600] and log CFU. Growth profile of wild-type
PezT is shown as (x) for A[600] values (on top) and black bars
in log CFU (bottom). Growth profiles of the following mutants
following IPTG induction are shown: T117V (  ; green bars), T120V (
 ;
blue bars), and K45A (  ; yellow bars). The
R157A, R170A, and D66T mutations yielded nontoxic growth
profiles similar to those of K45A and T117V and are not
presented in the figure.
|
|
|
|
|
|
The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
19606-19618)
copyright 2007.
|
|
|
|
|
|
|
