2 research outputs found
Crystal Structures of the F and pSLT Plasmid TraJ N‑Terminal Regions Reveal Similar Homodimeric PAS Folds with Functional Interchangeability
In the F family of conjugative plasmids,
TraJ is an essential transcriptional
activator of the <i>tra</i> operon that encodes most of
the proteins required for conjugation. Here we report for the first
time the X-ray crystal structures of the TraJ N-terminal domains from
the prototypic F plasmid (TraJ<sub>F</sub><sup>11–130</sup>) and from the Salmonella virulence
plasmid pSLT (TraJ<sub>pSLT</sub><sup>1–128</sup>). Both structures
contain similar Per-ARNT-Sim (PAS) folds, which further homodimerize
through the N-terminal helix and the structurally conserved β-sheet
of the PAS fold from each protomer. Mutational analysis reveals that
the observed dimeric interface is critical for TraJ<sub>F</sub> transcriptional
activation, indicating that dimerization of TraJ is required for its <i>in vivo</i> function. TraJ is specific in activating its cognate <i>tra</i> operon promoter; however, heterologous PAS domains from
pSLT and R100 TraJ can functionally replace the TraJ<sub>F</sub> PAS
domain, suggesting that the allelic specificity of TraJ is solely
mediated by the region C-terminal to the PAS domain
<i>Bacillus anthracis</i> Inosine 5′-Monophosphate Dehydrogenase in Action: The First Bacterial Series of Structures of Phosphate Ion‑, Substrate‑, and Product-Bound Complexes
Inosine 5′-monophosphate dehydrogenase (IMPDH)
catalyzes
the first unique step of the GMP branch of the purine nucleotide biosynthetic
pathway. This enzyme is found in organisms of all three kingdoms.
IMPDH inhibitors have broad clinical applications in cancer treatment,
as antiviral drugs and as immunosuppressants, and have also displayed
antibiotic activity. We have determined three crystal structures of <i>Bacillus anthracis</i> IMPDH, in a phosphate ion-bound (termed
“apo”) form and in complex with its substrate, inosine
5′-monophosphate (IMP), and product, xanthosine 5′-monophosphate
(XMP). This is the first example of a bacterial IMPDH in more than
one state from the same organism. Furthermore, for the first time
for a prokaryotic enzyme, the entire active site flap, containing
the conserved Arg-Tyr dyad, is clearly visible in the structure of
the apoenzyme. Kinetic parameters for the enzymatic reaction were
also determined, and the inhibitory effect of XMP and mycophenolic
acid (MPA) has been studied. In addition, the inhibitory potential
of two known <i>Cryptosporidium parvum</i> IMPDH inhibitors
was examined for the <i>B. anthracis</i> enzyme and compared
with those of three bacterial IMPDHs from <i>Campylobacter jejuni</i>, <i>Clostridium perfringens</i>, and <i>Vibrio cholerae</i>. The structures contribute to the characterization of the active
site and design of inhibitors that specifically target <i>B.
anthracis</i> and other microbial IMPDH enzymes