APPLICATION OF PLANT BIOTECHNOLOGY FOR INDUSTRIAL ENZYMES

Currently most infustrial enzymes are produced from microbes by large scale fermentation.
An alternative approach to the production of such enzymes would be to express and recover these enzymes from transgenic plant.
The production of industrial enzymes in transgenic alfalfa has certain advantages.
Alfalfa is widely grown perennial crop capable of three or more harvests in year.
It is beneficial environmentally since it does not require annual tilling and planting and needs fewer application of fertilizers and pesticides than conventional row crops.
Furthermore technology for extracting protien from alfaalfa while leaving avaluable residue for animal feed is well developed. Some enzymes and their applications are listed below:
a) α – amylase : Textiles, starch syrups, laundry and dish washing, detergents, paper designing, fermentation of ethanol, animalfeeds.
b) Cellulase : Dish washing detergents, animal feeds, textiles, bioenergy production.
c) Invertase : Manufacture of syrup from cane and beet sugar.
d) Lactase : Eliminates lactose from dairy food
e) Pectinase : fruit processing
f) Acid protienases : baking (improves dough handling)
g) Alkaline proteases : detergent, leather & fur.
h) Pepsin : cheese production
i) Subtilisin : chiral resolution of chemical compounds (or) pharmaceuticals
j) Lysozymes : anti bacterial
k) Isomerase : conversion of glucose to high fructose corn syrup
l) Alchohol dehydrogenase : chiral synthesis of chemicals
m) Peroxidase : laundry & wood pulp bleaches
n) Acetolactate decarboxilase : brewing industry
o) Histidine : cosmetics
p) Xylanase : paper industry

Manganese dependent lignin peroxidase from the fungus phanerochaete chrysosporium has been expressed in transgenic alfalfa plants.
This enzyme has potential for large scale industrial usage for lignin degradation and as a bleaching agent in bio-pulping processes (paper industry).
Expressing of this enzyme IN alfalfa may also have the potential for increasing the fibre degestability of forages for ruminants
α – amylase from Bacillus licheniformis has been expressed in alfalfa.
It was found to be a robust enzyme that is active over a wide pH and temperature range.
Transgenic alfalfa expressing the animal feed enzyme phytase (from Aspergillus niger) and two different cellulases have been field tested.
Phytase was expressed at levels upto 2% total soluble protein in some transgenic lines.
This makes it an economically viable alternative to microbially produced phytase.
The production of animal feed supplements in transgenic alfalfa is particularly appealing since the expressed juice or leaf meal preperation can be added to animal rations with no intervening extraction (or) purification neccesary.
They contain proteins and xanthophyll, a yellow pigment used to enhance the colour of poultry, making it more attractive to cusumers.
It has been recently dicovered that the cellulase enzyme can replace the pumice stones used in textile industry to produce stone wash denim. This will help to counter the damage that pumice stone can cause to fabric.
The cellulase enzyme can also be used as a biopolishing agentas it removes the fuzz from the surface of the cellulose fibres.
Proteases and hydrolases are used in laundry detergents and starch processing respectively.
Genetic manipulation can generate simple molecules from known chemical structures to ore active compounds.
For instance, sweetness of cornsyrup can be increased by chemical transformation using the glucose isomerase enzyme.
These developments may have very wide applications in pharmaceutical, food and agricultural areas.
Transgenic cotton plants with insect resistance have been produced by transferring a gene from the bacteria Bacillus thuringiensis. With this success, scientists are now trying to develop transgenic coloured cotton, which could replace the bleaching and dying process.
The manipulation of enzyme structure for having maximum catalytic activity and resistance to reaction environment is called enzyme engineering.
Many enzymes are sensitive to heat. Some enzymes catalyse reversible reactions. Some enzymes are sensitive to pH fluctuations.
The reaction kinetics are very poor while using such enzymes.
Genetic engineers have manipulated the genes coding for the enzymes in such a way as to have desired features.
More number of disulfide bonds in enzymes, increases the thermostability as well as resistance against extreme pH changes.
Nucleotides coding for cystine are introduced into the gene coding for the enzyme, by site directed mutagenesis.
The modified gene produces enzymes with some more cystine residues and establishes some additional disufide bonds.
The engineered enzymes are more stable at high temperature and extreme pH changes.
As the major portion of the enzymes remain the same, there is no change in the catalytic activity.
Another reason for poor stability towards high temperature is the presence of more asparagine in the enzyme molecule.
One or two asparagine residues are replaced by threonine or isoleucine by site ddirected mutagenesis in the gene coding for that enzyme.
The resulting enzymes are more stable towards heat