Neisseria meningitidis expresses a single 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase that is inhibited primarily by phenylalanine.

Neisseria meningitidis is the causative agent of meningitis and meningococcal septicemia is a major cause of disease worldwide, resulting in brain damage and hearing loss, and can be fatal in a large proportion of cases. The enzyme 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DAH7PS) catalyzes the first reaction in the shikimate pathway leading to the biosynthesis of aromatic metabolites including the aromatic acids l-Trp, l-Phe, and l-Tyr. This pathway is absent in humans, meaning that enzymes of the pathway are considered as potential candidates for therapeutic intervention.

Examining the role of intersubunit contacts in catalysis by 3-deoxy-d-manno-octulosonate 8-phosphate synthase.

3-Deoxy-d-manno-octulosonate 8-phosphate synthase (KDO8PS) catalyzes the reaction between phosphoenolpyruvate and arabinose 5-phosphate (A5P) in the first committed step in the pathway to 3-deoxy-d-manno-octulosonate, a component in the cell wall of Gram-negative bacteria. KDO8PS is evolutionarily and structurally related to the first enzyme of the shikimate pathway, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DAH7PS), which uses erythrose 4-phosphate in place of A5P. Both KDO8PS and type Iβ DAH7PS enzymes adopt similar homotetrameric associations with their active sites close to one of the interfaces.

Targeting the role of a key conserved motif for substrate selection and catalysis by 3-deoxy-D-manno-octulosonate 8-phosphate synthase.

3-Deoxy-D-manno-octulosonate 8-phosphate synthase (KDO8PS) catalyzes the reaction between three-carbon phosphoenolpyruvate (PEP) and five-carbon d-arabinose 5-phosphate (A5P), generating KDO8P, a key intermediate in the biosynthetic pathway to 3-deoxy-D-manno-octulosonate, a component of the lipopolysaccharide of the Gram-negative bacterial cell wall. Both metal-dependent and metal-independent forms of KDO8PS have been characterized. KDO8PS is evolutionarily and mechanistically related to the first enzyme of the shikimate pathway, the obligately divalent metal ion-dependent 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAH7PS) that couples PEP and four-carbon D-erythrose 4-phosphate (E4P) to give DAH7P.

Specificity and mutational analysis of the metal-dependent 3-deoxy-D-manno-octulosonate 8-phosphate synthase from Acidithiobacillus ferrooxidans.

3-deoxy-D-manno-octulosonate 8-phosphate synthase (KDO8PS) catalyzes the reaction between phosphoenol pyruvate and D-arabinose 5-phosphate to generate KDO8P. This reaction is part of the biosynthetic pathway to 3-deoxy-D-manno-octulosonate, a component of the lipopolysaccharide of the Gram-negative bacterial cell wall. Two distinct groups of KDO8PSs exist, differing by the absolute requirement of a divalent metal ion.

Reversing evolution: re-establishing obligate metal ion dependence in a metal-independent KDO8P synthase.

Two distinct groups of 3-deoxy-D-manno-octulosonate 8-phosphate synthase (KDO8PS), a key enzyme of cell-wall biosynthesis, differ by their requirement for a divalent metal ion for enzymatic activity. The unique difference between these groups is the replacement of the metal-binding Cys by Asn. Substitution of just this Asn for a Cys in metal-independent KDO8PS does not create the obligate metal-ion dependency of natural metal-dependent enzymes.

Arabinose 5-phosphate analogues as mechanistic probes for Neisseria meningitidis 3-deoxy-D-manno-octulosonate 8-phosphate synthase.

3-Deoxy-D-manno-octulosonate 8-phosphate (KDO8P) synthase catalyses the condensation reaction between phosphoenolpyruvate and D-arabinose 5-phosphate (D-A5P) in a key step in lipopolysaccharide biosynthesis in Gram-negative bacteria. The KDO8P synthase from Neisseria meningitidis was cloned into Escherichia coli, overexpressed and purified. A variety of D-A5P stereoisomers were tested as substrates, of these only D-A5P and l-X5P were substrates.

Reassessment of effects on lignification and vascular development in the irx4 Arabidopsis mutant.

The Arabidopsis thaliana irregular xylem4 (irx4) cinnamoyl-CoA reductase 1 (CCR1) mutant was reassessed for its purported exclusive rate-limiting or key effects on lignification. Analyses of gross growth characteristics and stem cross-section anatomy, from seedling emergence to senescence, revealed that stunted irx4 mutant lines were developmentally delayed, which in turn indirectly but predictably led to modest reductions (ca. 10-15%) in overall lignin amounts.

Characterization in vitro and in vivo of the putative multigene 4-coumarate:CoA ligase network in Arabidopsis: syringyl lignin and sinapate/sinapyl alcohol derivative formation.

A recent in silico analysis revealed that the Arabidopsis genome has 14 genes annotated as putative 4-coumarate:CoA ligase isoforms or homologues. Of these, 11 were selected for detailed functional analysis in vitro, using all known possible phenylpropanoid pathway intermediates (p-coumaric, caffeic, ferulic, 5-hydroxyferulic and sinapic acids), as well as cinnamic acid. Of the 11 recombinant proteins so obtained, four were catalytically active in vitro, with fairly broad substrate specificities, confirming that the 4CL gene family in Arabidopsis has only four members.

The Arabidopsis phenylalanine ammonia lyase gene family: kinetic characterization of the four PAL isoforms.

In Arabidopsis thaliana, four genes have been annotated as provisionally encoding PAL. In this study, recombinant native AtPAL1, 2, and 4 were demonstrated to be catalytically competent for l-phenylalanine deamination, whereas AtPAL3, obtained as a N-terminal His-tagged protein, was of very low activity and only detectable at high substrate concentrations. All four PALs displayed similar pH optima, but not temperature optima; AtPAL3 had a lower temperature optimum than the other three isoforms.

An in silico assessment of gene function and organization of the phenylpropanoid pathway metabolic networks in Arabidopsis thaliana and limitations thereof.

The Arabidopsis genome sequencing in 2000 gave to science the first blueprint of a vascular plant. Its successful completion also prompted the US National Science Foundation to launch the Arabidopsis 2010 initiative, the goal of which is to identify the function of each gene by 2010. In this study, an exhaustive analysis of The Institute for Genomic Research (TIGR) and The Arabidopsis Information Resource (TAIR) databases, together with all currently compiled EST sequence data, was carried out in order to determine to what extent the various metabolic networks from phenylalanine ammonia lyase (PAL) to the monolignols were organized and/or could be predicted.