The biochemical pathways leading to the production of valuable plant secondary metabolites are being studied with a virus to engineer increased production and to reduce cost of production. Over expression of the gene that encodes for the first enzyme (generally the rate limiting enzyme) in a pathway results in higher levels of the desired end product. This has been successfully done in the enhancement of taxol production from transformed tissue cultures of Taxus species with taxidine synthase gene.
The biosynthetic pathways for the production of plant phenolics are relatively well worked out.
Plant phenolics are a large variety of secondary metabolites that contain a phenol group, a hydroxyl functional group on an aromatic ring as shown below.
Most plant phenolics are synthesized through shikonic acid pathway and phenyl propanoid pathway.
The shikonic acid pathway converts simple carbohydrate precursors derived from glycolysis and the pentose phosphate pathway to the aromatic amino acids, which act as the precursors for the synthesis of phenolics as briefly given in the following
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To enhance phenylalanine synthesis it is essential to knockout any feedback inhibition of the pathway from chorismate to phenylalanine.
The next step was to remove competing pathways i.e the synthesis of tyrosine and tryptophan.
Once all control circuits and competing pathways are removed, attempts were made to increase the carbon flux through the biosynthetic pathway.
Over expressing all the gene in the pathway did not enhance the yield of phenylalanine because the supply of the precursor (E-4P and/or PEP) was rate limiting.
The precursors were increased by cloning transketolase (to enhance E4P levels) and eliminating pyruvate kinase (to enhance PEP levels).
The most abundant class of secondary phenolic compounds in plants are derived from phenylalanine via the elimination of an ammonia molecule to form cinnamic acid.
This reaction is catalyzed by phenylalanine ammonis lyase (PAL). This is perhaps the most studied enzyme in plant secondary metabolites and is situated at a branch point between primary & secondary metabolism, so the reaction that it catalyses is an important regulatory step in the formation of many phenolic compounds.
Reactions subsequent to that catalyzed by PAL lead to the addition of more hydroxyl groups and other substituent’s.
Trans – cinnamic acid, P – coumaric acid and other derivatives are simple phenolic compounds called phenylpropanoids, that form the important building blocks of the more complex phenolic compounds.
The phenyl propanoid pathway is clearly explained in the below diagram :
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The enzyme PAL is the first committed step for phenyl propanoid pathway.
This enzyme has been over expressed in tobacco plants and the transgenic plants showed resistance to pathogens like viruses, bacteria, fungi and grazing insect larvae. While suppression of PAL reduced resistance
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