3 research outputs found
Characterization of Two Late-Stage Enzymes Involved in Fosfomycin Biosynthesis in Pseudomonads
The
broad-spectrum phosphonate antibiotic fosfomycin is currently
in use for clinical treatment of infections caused by both Gram-positive
and Gram-negative uropathogens. The antibiotic is biosynthesized by
various streptomycetes, as well as by pseudomonads. Notably, the biosynthetic
strategies used by the two genera share only two steps: the first
step in which primary metabolite phosphoenolpyruvate (PEP) is converted
to phosphonopyruvate (PnPy) and the terminal step in which 2-hydroxypropylphosphonate
(2-HPP) is converted to fosfomycin. Otherwise, distinct enzymatic
paths are employed. Here, we biochemically confirm the last two steps
in the fosfomycin biosynthetic pathway of <i>Pseudomonas syringae</i> PB-5123, showing that Psf3 performs the reduction of 2-oxopropylphosphonate
(2-OPP) to (<i>S</i>)-2-HPP, followed by the Psf4-catalyzed
epoxidation of (<i>S</i>)-2-HPP to fosfomycin. Psf4 can
also accept (<i>R</i>)-2-HPP as a substrate but instead
performs an oxidation to make 2-OPP. We show that the combined activities
of Psf3 and Psf4 can be used to convert racemic 2-HPP to fosfomycin
in an enantioconvergent process. X-ray structures of each enzyme with
bound substrates provide insights into the stereospecificity of each
conversion. These studies shed light on the reaction mechanisms of
the two terminal enzymes in a distinct pathway employed by pseudomonads
for the production of a potent antimicrobial agent
Evaluation of a Cyclopentane-Based γ‑Amino Acid for the Ability to Promote α/γ-Peptide Secondary Structure
We report the asymmetric synthesis
of the γ-amino acid (1<i>R</i>,2<i>R</i>)-2-aminomethyl-1-cyclopentane carboxylic
acid (AMCP) and an evaluation of this residue’s potential to
promote secondary structure in α/γ-peptides. Simulated
annealing calculations using NMR-derived distance restraints obtained
for α/γ-peptides in chloroform reveal that AMCP-containing
oligomers are conformationally flexible. However, additional evidence
suggests that an internally hydrogen-bonded helical conformation is
partially populated in solution. From these data, we propose characteristic
NOE patterns for the formation of the α/γ-peptide 12/10-helix
and discuss the apparent conformational frustration of AMCP-containing
oligomers
Evaluation of a Cyclopentane-Based γ‑Amino Acid for the Ability to Promote α/γ-Peptide Secondary Structure
We report the asymmetric synthesis
of the γ-amino acid (1<i>R</i>,2<i>R</i>)-2-aminomethyl-1-cyclopentane carboxylic
acid (AMCP) and an evaluation of this residue’s potential to
promote secondary structure in α/γ-peptides. Simulated
annealing calculations using NMR-derived distance restraints obtained
for α/γ-peptides in chloroform reveal that AMCP-containing
oligomers are conformationally flexible. However, additional evidence
suggests that an internally hydrogen-bonded helical conformation is
partially populated in solution. From these data, we propose characteristic
NOE patterns for the formation of the α/γ-peptide 12/10-helix
and discuss the apparent conformational frustration of AMCP-containing
oligomers