Identification, characterization and functional expression of a tyrosine ammonia-lyase and its mutants from the photosynthetic bacterium Rhodobacter sphaeroides.

A tyrosine ammonia-lyase (TAL) enzyme from the photosynthetic bacterium Rhodobacter sphaeroides (RsTAL) was identified, cloned and functionally expressed in Escherichia coli, where conversion of tyrosine to p-hydroxycinnamic acid (pHCA) was demonstrated. The RsTAL enzyme is implicated in production of pHCA, which serves as the cofactor for synthesis of the photoactive yellow protein (PYP) in photosynthetic bacteria. The wild type RsTAL enzyme, while accepting both tyrosine and phenylalanine as substrate, prefers tyrosine, but a serendipitous RsTAL mutant identified during PCR amplification of the RsTAL gene, demonstrates much higher preference for phenylalanine as substrate and deaminates it to produces cinnamic acid.

Calcium alginate bead immobilization of cells containing tyrosine ammonia lyase activity for use in the production of p-hydroxycinnamic acid.

An Escherichia coli catalyst with tyrosine ammonia lyase activity (TAL) has been stabilized for repeated use in batch conversions of high tyrosine solids to p-hydroxycinnamic acid (pHCA). The TAL biocatalyst was stabilized by controlling the reaction pH to 9.8 +/- 0.

Development of a combined biological and chemical process for production of industrial aromatics from renewable resources.

Production of industrial aromatic chemicals from renewable resources could provide a competitive alternative to traditional chemical synthesis routes. This review describes the engineering of microorganisms for the production of p-hydroxycinnamic acid (pHCA) and p-hydroxystyrene (pHS) from glucose. The initial process concept was demonstrated using a tyrosine-producing Escherichia coli strain that overexpressed both fungal phenylalanine/tyrosine ammonia lyase (PAL) and bacterial pHCA decarboxylase (pdc) genes.

Functional expression of prokaryotic and eukaryotic genes in Escherichia coli for conversion of glucose to p-hydroxystyrene.

The chemical monomer p-hydroxystyrene (pHS) is used for producing a number of important industrial polymers from petroleum-based feedstocks. In an alternative approach, the microbial production of pHS can be envisioned by linking together a number of different metabolic pathways, of which those based on using glucose for carbon and energy are currently the most economical. The biological process conserves petroleum when glucose is converted to the aromatic amino acid L-tyrosine, which is deaminated by a tyrosine/phenylalanine ammonia-lyase (PAL/TAL) enzyme to yield p-hydroxycinnamic acid (pHCA).

Production of tyrosine from sucrose or glucose achieved by rapid genetic changes to phenylalanine-producing Escherichia coli strains.

Escherichia coli K12 strains producing L-phenylalanine were converted to L-tyrosine-producing strains using a novel genetic method for gene replacement. We deleted a region of the E. coli K12 chromosome including the pheA gene encoding chorismate mutase/prephenate dehydratase, its leader peptide (pheL), and its promoter using a new polymerase chain reaction-based method that does not leave a chromosomal scar.

Production of p-hydroxycinnamic acid from glucose in Saccharomyces cerevisiae and Escherichia coli by expression of heterologous genes from plants and fungi.

Biological production of p-hydroxycinnamic acid (pHCA) from glucose can be achieved via deamination of the aromatic amino acids l-tyrosine or l-phenylalanine. Deamination of l-phenylalanine produces trans-cinnamic acid (CA) which is further hydroxylated in the para position to produce pHCA. However, when tyrosine is used as the substrate, trans-pHCA is produced in one step.

Characterization of the Escherichia coli AaeAB efflux pump: a metabolic relief valve?

Treatment of Escherichia coli with p-hydroxybenzoic acid (pHBA) resulted in upregulation of yhcP, encoding a protein of the putative efflux protein family. Also upregulated were the adjacent genes yhcQ, encoding a protein of the membrane fusion protein family, and yhcR, encoding a small protein without a known or suggested function. The function of the upstream, divergently transcribed gene yhcS, encoding a regulatory protein of the LysR family, in regulating expression of yhcRQP was shown.

Crystal structure of phenylalanine ammonia lyase: multiple helix dipoles implicated in catalysis.

The first three-dimensional structure of phenylalanine ammonia lyase (PAL) has been determined at 2.1 A resolution for PAL from Rhodosporidium toruloides. The enzyme is structurally similar to the mechanistically related histidine ammonia lyase (HAL), with PAL having an additional approximately 160 residues extending from the common fold.