INDUSTRIAL BIOTECHNOLOGY:

MICROBIAL BIOTECHNOLOGY
PLANTS AS BIOREACTORS

NO. 12

MICROBIAL BIOTECHNOLOGY
Microbes (or microorganisms) are minute living things that individually are too small to
be seen with the naked eye. Microorganisms are a diverse and unique life form. They
include bacteria, fungi (yeasts and molds), protozoa, microalgae, and viruses. This
group of organisms is usually associated only with uncomfortable infections, major
diseases such as AIDS or such common inconveniences as spoiled food. Microbes live in
familiar settings such as soil, water, food, and animal intestines, as well as in more
extreme settings such as rocks, glaciers, hot springs, and deep-sea vents. The wide
variety of microbial habitats reflects an enormous diversity of biochemical and
metabolic traits that have arisen by genetic variation and natural selection in microbial
populations.

Historically, humans have exploited some of this microbial diversity in the production of
fermented foods such as bread, cheese, yoghurt, alcoholic beverages like wine and beer
and condiments like vinegar, fish sauce and soy sauce. Some soil microbes release
nitrogen that plants need for growth by breaking down waste and incorporating
nitrogen gas into the organic compounds thereby recycling and maintaining the critical
composition of the Earth's soil, water and atmosphere. Other microbes challenge the
food supply by causing yield-reducing diseases in food-producing plants and animals. In
our bodies, different microbes help to digest food, synthesize some vitamins, ward off
invasive organisms, and engage in skirmishes and pitched battles with the human
immune system in the give-and-take of the natural disease process. In general,
microorganisms make crucial contributions to the welfare of the world's inhabitants by
helping to maintain the balance of living organisms and chemicals in our environment.

The microorganisms' most notable characteristics: simple structures, rapid

reproduction, and low cost of growth and maintenance, have made them extremely
useful as biological and biotechnological research models and tools. Likewise, with the
advent of modern biotechnology and continuously advancing research procedures, the
simple characteristics and ease of handling microorganisms made them of commercial
and economic significance in the developed as well as developing countries. Commercial
and environmental application of microorganisms assures human, especially the next
generation, high-quality food and a healthy and safe Earth to live.
Microbial biotechnology is the application of scientific and engineering principles to the
processing of materials by microorganisms (such as bacteria, fungi, algae, protozoa and
viruses) to create useful products or processes. The microorganisms utilized may be
natural isolates, laboratory selected mutants or microbes that have been genetically
engineered using recombinant DNA methods. For example, most antibiotics come from
microbial fermentations involving a group of organisms called actinomycetes. Yeasts are
used in baking, in the production of alcohol for beverages and in fuel production
(biofuel). Other groups of microorganism form products that range from organic acids
(citric acid, lactic acid) to enzymes (carbohydrases, proteases, lipases) used for the
production of various carbohydrates, amino acids and detergents. For example,
aspartame, a sweetener, is derived from amino acids produced by microorganisms.
Microbial biotechnology also deals with the prevention of deterioration of processed or
manufactured goods environmental protection and with waste disposal systems. In
addition, more and more improved vaccines and better disease-diagnostic tools,
development of new industrial catalysts and development of new microbial agents for
bioremediation of soil and water contaminated by agricultural runoff will continuously
be available for human to consume and to depend on.

Microbial Biotechnology in Foods and Agriculture

The food and agricultural industries are among the key areas targeted by biotechnology.
Some of the benefits that advanced techniques of biotechnology are capable of
conferring on food and agricultural industries include the following:

1. Development of genetically engineered plants which have internal resistance to

drought, frost, insect pests and infestation;

2. Reduction in dependency of plants on chemical fertilizers and identification of

alternatives to expensive fertilizers such as nitrogen fertilizers which require very
large amounts of energy for their production;

3. Replacement of dangerous chemical pesticides with bio-pesticides (microbial

pesticides) to manage and control the problem of pests;

4. Reduction in the reliance on chemical treatments to control weeds by engineering

herbicide tolerance into crops;

5. Production of products that have high yield and enhanced nutritional value; and

6. Development of novel biomass products as foodstuffs, using organisms such as

algae, fungi, bacteria and yeast
 Food
Fermentation is the process of bioconversion of organic substances by microorganisms
and/or enzymes (complex proteins) of microbial, plant or animal origin. It is one of the
oldest forms of food preservation, which is applied globally. Indigenous fermented foods
such as bread, cheese and wine, have been prepared and consumed for thousands of
years and are strongly linked to culture and tradition, especially in rural households and
village communities (Achinewu et al., 1991, 1992). It is estimated that fermented foods
contribute to about one third of the diet worldwide (FAO, 2004).

During fermentation processes, microbial growth and metabolism (the biochemical

processes whereby complex substances and food are broken down into simple
substances) result in the production of a diversity of metabolites which include enzymes
capable of breaking down carbohydrates, proteins and lipids present within the
substrate and/or fermentation medium; vitamins; antimicrobial compounds (e.g.
bacteriocins and lysozyme), texture-forming agents (e.g. xanthan gum), amino acids,
glutamic acids, organic acids and flavor compounds (e.g. esters and aldehydes). Many of
these microbial metabolites are produced at the industrial level in both developed and
developing countries for use in food processing applications.

Biotechnology research incorporates the use of genetic technologies for strain

development and improvement, and for diagnostic studies (Tietyen et al., 2000; FAO,
2004). While microorganisms are beneficial in most fermentation processes, some may
pose the risk of food contamination and can cause food-borne illness. Diagnostic
methodologies which integrate the use of molecular genetic techniques enhance the
speed and sensitivity of microbial testing and are increasingly being applied in
developing countries (FAO, 2000, 2004).

Genetically modified (GM) wine has been marketed in the United States for the past two
years (Cummin, 2005). Presently, only one modification has been acknowledged but
there are a number of modifications reported in wine yeast and bacteria and a number
of modifications reported for grapes. The United States Food and Drug Administration
(FDA) in 2003 designated the yeast, Saccharomyces cereviseae strain ML01 to be a
substance generally recognized as safe (GRAS). The strain included a gene for malolactic
enzyme from the bacterium Oenococcus oeni and a malate permease gene from the
fission yeast Schizosaccharomyces pombe. Wine making consists of alcoholic
fermentation employing the metabolic pathways of yeast and malolactic fermentation to
convert malic acid to lactic acid to reduce the acidity of the wine. Malolactic acid
fermentation is usually achieved using lactic acid bacteria which have a permease for
malic acid. Having the two fermentations in one organism spares the need for the lactic
acid fermentation. The recombinant yeast softened the wine’s mouth feel by decreasing
its acidity. Its use also reduces buttery flavors (diactyl) due to lactic acid secondary
metabolism. The company distributing the GM yeast is Springer Oenologie, Lesaffre
Group, of North America.

In 1990, the United Kingdom became the first country to permit the use of a live,
genetically-modified organism in food. This was a special strain of bakers' yeast
engineered to make the bread dough rise faster. Existing genes were placed under the
control of stronger, constitutive promoters, which help the yeast to break down the
sugar maltose faster than usual. This yeast was and is not used by any food
manufacturers, however.

Molecular diagnostics provide outstanding tools for the detection, identification and
characterization of microbial strains for bio-processing applications and for the
improvement of fermentation processes. Many of the classical food microbiological
methods used in the past were culture-based, with microorganisms grown on agar plates
and detected through biochemical identification. These methods are often tedious, labor
intensive and slow. Genetic based diagnostic and identification systems can greatly
enhance the specificity, sensitivity and speed of microbial testing. Molecular
methodologies, including polymerase chain reaction (PCR) and ribotyping, can be used
for typing microbial strains, for characterizing and monitoring the presence of improved
microbial strains as well as detection of spoilage microflora (microbes causing food to
become unfit for eating), food chain or to identify the causal agents of food borne
illnesses. Monoclonal and polyclonal antibodies can also be used for diagnostics, e.g. in
enzyme-linked immunosorbent assay (ELISA) kits (FAO, 2000, 2004). Microarrays are
also referred to as biochip, DNA chip, DNA microarray or gene arrays and offer
unprecedented opportunities and approaches to diagnostic and detection methods. They
can be used for the detection of pathogens, pesticides and toxins and offer considerable
potential for facilitating process control, the control of fermentation processes and
monitoring the quality and safety of raw materials (FAO, 2000, 2004; BREI, 2006).

Genomics
In recent years, the genome sequences of many food-related microorganisms have been
completed (e.g. S. cerevisiae, commonly known as baker's or brewer's yeast, was the
first eukaryote to have its genome sequenced - in 1996) and large numbers of microbial
genome sequencing projects are also underway. Functional genomics, a relatively new
area of research, aims to determine patterns of gene expression and interaction in the
genome, based on the knowledge of extensive or complete genomic sequence of an
organism. It can provide an understanding of how microorganisms respond to
environmental influences by expressing specific genes in different situations or
ecologies. For a range of microorganisms, it is now possible to observe the expression of
many genes simultaneously, even those with unknown biological functions, as they are
switched on and off during normal development or while an organism attempts to cope
with pathogens or changing environmental conditions.

Agriculture
Applications of biotechnology to plants or animals have improved their food processing
properties and the production of proteins from genetically modified (GM)
microorganisms to improve plant or animal production and a hormone- increasing milk
production in dairy cows using GM bacteria (Lemaux, 2000; Tietyen et al., 2000).
Scientists are not able to exchange genetic information between bacteria, plants, and
animals (including humans). These new techniques have prompted considerable debate
on the ethical and moral aspects of this branch of science.

Also, Upjohn Company of Kalamazoo, Michigan developed a squash line (called ZW 20)
that contains the coat protein genes of watermelon mosaic virus 2 (WMV2) and zucchini
yellow mosaic virus (ZYMV). The modifications have demonstrated remarkable field
resistance against the two viruses.

Furthermore, to combat grapevine fanleaf virus (GFLV), scientists at LVMH, Inc., in

Paris, France have used a bacterium vector to introduce a GFLV coat protein gene into a
chardonnay grape variety. According to LVMH, Inc., the gene has been effectively
transferred and the engineered grapevines have exhibited resistance to infection by the
virus.

To control weeds, chemical herbicides have come to play a significant role in agriculture.
However, there are risks associated with the use of chemical herbicides. In addition to
causing serious environmental problems due to chemical contaminants, chemical
herbicides themselves have undesirable effects on non-target organisms. With the help
of rDNA technology, scientists are hoping to genetically modify plants tolerant to
herbicides. Herbicide tolerance can occur when the phytotoxic compound fails to be
taken up by living tissue or is rendered non-phytotoxic by conjugation, hydrolysis, or
another metabolic event (detoxification).

By using rDNA technology, the genes that code for the protein involved can be
identified, isolated and modified by directed mutagenesis and introduced into plant
cultivar to confer the herbicide-tolerant phenotype. To reduce dependency on chemical
fertilizers, agricultural biotechnologists are conducting considerable research using
genetic manipulation to increase the range of plants that can fix atmospheric nitrogen.
Microbial Biotechnology in Chemical Industries
Chemical industries are involved in the production of specialty chemicals such as amino
acids, enzymes, polysaccharides, vitamins, sweeteners, food additives, flavors,
fragrances, etc. These industries are also interested in converting biomass to produce
specialty chemicals from either plants or biological wastes, such as those generated from
agriculture and food processing. Although advanced techniques of biotechnology have
not yet played a significant role in chemical production, there are areas of chemical
industries, however, where this technology can have substantial impact, such as the
production of amino acids, enzymes and polysaccharides. Manufacturers are
particularly interested in the potential for producing existing and new chemicals at
lower cost with reduced energy requirements and waste disposal problems. Biocatalytic
chemical production has the added advantage of specificity, controllability, low
temperature operation, environmental acceptability and simplicity. For example, much
of the present organic chemical industry is based upon petroleum and most of the
chemical intermediates generated are partial oxidation products. Specific, controlled,
partial oxidation is difficult to achieve by conventional catalysis. By using
microorganisms, this type of reaction can be easily realized.

Amino Acids
Amino acids are the building blocks of proteins in animals, plants and microorganisms.
They can be produced either by isolation from natural materials – from hydrolysis of
plant proteins – or by chemical, microbial or enzymatic synthesis. Whereas chemical
synthesis produces a racemic (optically inactive) product that may require additional
resolution, microbial and enzymatic syntheses give rise to optically pure amino acids.
Commercially, amino acid producing bacteria have been used since the 1950s. With the
help of biotechnology, strains have been subsequently improved genetically by the
generation of auxotroph or regulatory mutants.

Enzymes
Industrial applications of enzymes include the production of cheese, the clarification of
apple juice, the development of more efficient laundry detergents, pulp and paper
production and the treatment of sewage. These processes have been dramatically
enhanced by the use of recombinant DNA techniques to design enzymes of increased
activity, stability and specificity. Enzymes are biological catalysts used to facilitate and
speed up metabolic reactions in living organisms. They are proteins and require a
specific substrate on which to work. Their catalyzing conditions are set within narrow
limits, e.g. optimum temperature, pH conditions and oxygen concentration. Most
enzymes are denatured at temperatures above 42°C. However, certain bacterial enzymes
are tolerant to a broader temperature range. Enzymes are essential in the metabolism of
all living organisms and are widely applied as processing aids in the food and beverage
industry (FAO, 2000, 2004; BREI, 2006).

Protein Engineering
Using modern techniques of biotechnology, scientists are attempting to improve the
yield of an enzyme by transferring the encoding gene to a microorganism capable of
producing the enzyme in larger amounts. Because they are large and fragile molecules,
enzymes tend to change their nature when exposed to heat, solvents and other extreme
conditions that characterize most industrial processes. To remedy these problems, the
current research seeks to modify the genetic information coding for an enzyme in order
to introduce new chemical properties into the molecule, such as new chemical bonds
that stabilize its structure. Enzymes, of (hyper) thermophilic, psychrophilic and/or
extremophilic origin as well as those engineered and/or evolved to exhibit function even
in a very unusual environment as currently being explored for understanding of the
structure and function relationship for proteins and also for its biotechnological
potential.

In the past, enzymes were isolated primarily from plant and animal sources, and thus a
relatively limited number of enzymes were available to the food processor at a high cost.
Today, bacteria and fungi are exploited and used for the commercial production of a
diversity of enzymes. Several strains of microorganisms have been selected or
genetically modified to increase the efficiency with which they produce enzymes. In
most cases, the modified genes are of microbial origin, although they may also come
from different kingdoms. For example, the DNA coding for chymosin, an enzyme found
in the stomach of calves, that causes milk to curdle during the production of cheese, has
been successfully cloned into yeasts (Kluyveromyces lactis), bacteria (E. coli) and molds
(Aspergillus niger var. awamori). Chymosin produced by these recombinant
microorganisms is currently commercially produced and is widely used in cheese
manufacture (FAO, 2000, 2004).

Directed evolution is one of the main methods currently used for protein engineering.
This technique involves creating large numbers of new enzyme variants by random
genetic mutation and subsequently screening them to identify the improved variants.
This process is carried out repeatedly, thus mimicking natural evolution processes
(Arnold, 1996; Tietyen et al., 2000; IFIC, 2000; Biotech, 2000; ADA, 2000; Arnold et
al., 2001).
Polysaccharide
Polysaccharide has important application in the food, cosmetics, chemical, medical and
oil industries. Polysaccharides are produced by yeast, fungi and bacteria. They are also
naturally available as cellulose, lignin and chitin. Polysaccharides are used as lubricants,
viscosifiers, flocculating and gelling agents in food processing, and for stabilizing liquid
suspensions. At present, significant research is being conducted to apply modern
techniques of biotechnology to the production of microbial polysaccharide. Using
advanced biotechnology, current research is also directed toward the conversion of
biomass feedstocks to fermentable substrates, such as cellulose to glucose. While this
generally requires relatively expensive microbial enzymes (cellulose) possessing
desirable characteristics such as thermostability and high activity, manufacturers are
also able to combine the desirable characteristics of several less optimal cellulose
enzyme producers using recombinant DNA technology. Many laboratories are involved
in detection, identification, isolation and gene transfer for strain improvement.

Microbial Biotechnology in Medicine and Pharmaceutical

Industries
Insulin

In the fields of medicine and pharmacology, the initial application of advanced

biotechnology has been in the production of human insulin for the treatment of
diabetes.

Vaccines

The idea behind vaccination as a means of immunization is that by injecting a weakened

or dead pathogenic microbe into an individual, the body recognizes the antigen as
foreign and stimulates antibodies to destroy the invading organisms. It is not always
possible for vaccines to be completely risk free. Since the entire genetic material of the
pathogen is contained in vaccines, occasionally they cause the infectious disease that
they intend to cure. Using rDNA technology, scientists are trying to produce synthetic
vaccines, which use only a part of a specific antigen of the pathogenic organism, thereby
eliminating the disease causing capability of vaccines.
Interferon

Interferons are a family of proteins that have two important biological effects –
inhibition of cellular proliferation (potentially with anti-cancer action) and modulation
of the immune system. Prior to using rDNA technology, interferon was obtained from
human white blood cells. With the help of rDNA technology, “a human leucocyte
interferon (gene 514) base pair long ago was synthesized, incorporated into a plasmid,
and subsequently cloned into E. coli.” It is interesting to note that while natural
interferon is a glycosylated protein, the interferon produced by rDNA technology is not.

DNA Probes

Using “restriction enzymes” scientists are able to cut DNA into fragments and devise
DNA probes, and thereby identify defects and mutations that can cause diseases. DNA
probes have been used in the diagnosis of the bacteria that cause periodontal (gum)
disease, and a variety of genetic diseases such as Duchenne muscular dystrophy, cystic
fibrosis and Huntington's disease.

Gene Therapy

Both natural and chemically enhanced microbial products can be used to control
human, animal and plant diseases. Using traditional genetics or recombinant DNA
techniques, the microorganism can be modified to improve the yield or action of
antibiotics and other antimicrobial agents. New research directions are aimed at
discovering microbial metabolites with pharmacological activities useful in the
treatment of hypertension, obesity, coronary heart disease, cancer and inflammation.
Vaccines are essential to protect humans and animals from microbial diseases.
Recombinant DNA technology has allowed the production of novel vaccines that offer
protection without the risk of infection (e.g. hepatitis B vaccine). Other examples
include the use of microbial cells to produce human or animal biologicals such as
insulin, growth hormone and antibodies. The industrial microbiologist/biotechnologist
may screen new microbial sources (e.g., marine microorganisms) for their ability to
produce new pharmaceuticals.
Microbial Biotechnology in the Industries

Energy

Biotechnology can play an important role in the production of fuels from organic matter
via biomass conversion. The most prominent fuel presently, petroleum, is a
nonrenewable source that poses environmental risks in its extraction and use.
Biotechnology, by using available and abundant sources of biomass, could generate a
renewable and less environmentally hazardous source of energy called bio-energy. Bio-
energy, in the form of methane gas, may also be produced by bacteria consuming sewage
sludge in anaerobic conditions. Modern techniques of biotechnology can also be used to
enhance oil recovery. It is believed that conventional oil-extracting technologies are
capable of extracting only 50% of the world's oil supply, while the remaining 50% is
trapped in rock or is too thick to pump.

Mining

The environmental and health hazards associated with traditional mining technologies
have led many mining industries in the past few years to turn to a more efficient and
environmentally risk-free method for extracting minerals from ores that use micro-
organisms to leach metals from mine dumps. Microbial mining has improved recovery
rates and reduced costs. The iron-oxidizing bacterium Thiobacillus ferooxidans has
been used for bacterium-catalyzed leaching.

Microbial Biotechnology in the Environment

The application of biotechnology in the area of environmental protection is not a new
phenomenon – for example, microorganisms have long been used in sewage treatment
to eliminate solid particles. It was in the early part of the 20th century that an activated
sludge process for mineralizing organic waste was first developed. Since then, this
process has become more complex and sophisticated, incorporating many modern
technologies. Meanwhile, the use of anaerobic digestion processes for the treatment of
wastes, and the coincidental production of biogas (mainly methane and CO2), has
become an important source of energy generation. It is believed that recent
development in biotechnology could play an increasingly important role in pollution
prevention and toxic waste treatment via bioremediation, biotreatment and
biofiltration.
Bioremediation

Bioremediation, also referred to as biodegradation, is the use of organisms to degrade

waste materials into less toxic or non-toxic material in the environment. Detoxification
is when the waste is made less toxic. Mineralization is when the waste material is
converted into inorganic compounds (CO2, H2O, CH3) (Martello, 1991). Microbes and
other organisms use organic substances for nutrients and energy. Bioremediation is
simply these organisms using “waste products” as a nutrient source. This is not a new
concept. Composting is a form of bioremediation where scraps from the kitchen or
garden are left outside to be broken down so the remaining nutrients can be returned to
the soil.

In 1989, bioremediation with naturally occurring microbes was used to aid in the
cleanup of a massive oil spill in Alaska. The Exxon Valdez oil tanker spilled
approximately 11 million gallons of crude oil that contaminated over 1,000 miles of
shoreline (Princeton, 1990). To encourage the oil-degrading bacteria, nitrogen and
phosphorous were sprayed over the beaches for an extended period of time. Several
months later, Exxon and the EPA had treated 70 million miles of coastline (Princeton,
1990). In a study by the National Marine Fisheries Service, not only did they find oil
from the spill still present on some of the beaches, but also that it was enough to still be
harmful to the native wildlife (Rosen, 2003). These data disprove the presumed
successful outcome of the bioremediation efforts of the Exxon Valdez spill.

Through genetic engineering, scientists believe they can increase the effectiveness of
bioremediation. With biotechnology, bioremediation has great potential. Microbes can
be modified to be able to survive and degrade toxins and pollutants that would normally
be fatal or impossible to break down.

Waste and wastewater management

Microbial communities can detoxify contaminants in water, soils, sediment, and sludge.
This allows society to reclaim their resource value. Microbial communities can convert
the energy value in various types of biomass from its diffuse and sometimes hazardous
form to energy outputs that are readily used by human society: e.g., methane, hydrogen,
electricity, ethanol, and biodiesel.

As encountered above, there are many bacteria that can break down different wastes.
However, much of the problem remains in the bacteria not being able to survive in the
environment where the waste is located. Aquatic bacteria may not be able to live in soil,
soil dwelling bacteria may not be able to live in the water, warmer climate bacteria may
not be able to flourish enough in cold environments to break down waste. All these are
obstacles that may be eluded by genetic engineering.

There is a great deal of benefits to bioremediation. The microorganisms used in

bioremediation have been modified to exhibit higher metabolisms, and therefore
detoxify the site faster than naturally occurring organisms could. The cost of
development and implication is far less than traditional techniques. The contaminants
are broken down into harmless components, so the pollution is eliminated, making the
treatment permanent. In many traditional methods of cleanup the toxins are simply
removed from the site, which cause a great deal of disturbance to the environment
(Martello, 1991; Heaf, 2000).

GM bioremediation is still in the experimental stage, and has not yet been approved for
use in the environment. This is for fear of the negative aspects listed above coming true.
Once more studies have been done, and we have a better understanding of how these
organisms will behave in the environment, bioremediation may prove be our best
alternative. Future possibilities are endless; problems concerning radioactive waste
disposal may be obsolete, land that has been rendered unviable could be restored, and
lakes, streams, and rivers could be completely decontaminated. There is a lot of
potential for a safer, healthier, cleaner environment through GM bioremediation.

PLANTS AS BIOREACTORS
Industrially important enzymes are not only produced by microorganisms. There are
plant proteins that are being considered for industrial applications. Higher plants,
however, are usually not regarded as prolific producers of many commercially important
proteins because many industrially important proteins synthesized in plants are also
found in other biological sources and other sources becomes choice for both technical
and economic reasons. Plant growth is seasonal in nature and hence a constant source
of material is not always possible. Higher plants also tend to accumulate waste
substances in structures called vacuoles. Upon cell disruption of these wastes, which
include a number powerful precipitating and denaturing agents, often irreversibly
inactivate many plant proteins
Examples of Plant Proteins for Industry
Monellin and thaumatin – two plant proteins recognized as the sweetest-known
naturally occurring substances
-amylases (like barley) play important role in starch processing industry
Papain – proteolytic enzyme (also known as the vegetable pepsin from papaya
used as meat tenderizer, used for batting of animal skin for the leather industry,
digestive aid, and debriding agent (for cleaning of wounds)
Ficin – commercially available protease (used in food processing industry)

Why Use Plants As Bioreactors

Production of antibodies and antibody segments in transgenic plants
It is believed to result in stable integration of the foreign DNA into plant genome
unlike in microbe where foreign genes are plasmid encoded and may be lost
Processing and assembly of foreign proteins in plants (Table 7.1) are similar to
those in animal cells (including posttranslational modification)
Plants are inexpensive to grow on large scale and production not limited by
fermentation capability
Since most foreign genes might be produced in seeds that will be stable and will
be maintained for long periods under ambient conditions
Polymer is less costly if produced in plants
Foreign proteins in seeds (oil body proteins) ease of purification and high rate of
production

Production of Foreign Proteins in Plants

Studies have been conducted to introduce gene from different sources into plants (Table
7.1 and 7.2). To date expression of foreign proteins in genetically modified plants are
relatively low and attempts to address this problem is still a subject of research because
of the potential for using plants as bioreactors for foreign proteins.
Table 7-1. Proteins produced in plants. List of known proteins
from different sources (humans, bacteria, fungi, etc.) expressed
in host plants (tobacco, potato,etc.)
Protein Source Host plant Expression in plant host

Alpha amylase bacteria tobacco 0.3% of soluble leaf protein

Chymosin
Calf tobacco 0.5% of soluble proteins

Cyclodextrin
Bacteria potato 0.01% of soluble tuber
Glycosyltransferase

human 0.003% of soluble leaf

Erythropoietin tobacco
proteins

Glucoamylase Fungi potato not reported

Growth hormone Trout tobacco 0.1 % of soluble leaf protein

0.007% of soluble leaf

Hepatitis B surface Virus tobacco
protein

Hirudin Leech canola % of seed wt

Interferon B Human tobacco 0.00002% of fresh wt

Lysozyme Chicken tobacco 0.003 % leaf tissue

tobacco
Phytase Fungi 14.4% soluble leaf protein

human 0.02% of soluble leaf

Serum almbumin potato
protein

tobacco
Xylanase Bacteria 4.1 % of soluble leaf protein
Table 7-2. Plants as bioreactors. Advantages and disadvantages of
plants used as bioreactors in industry.
Advantages
Likely economically attractive production
cost
Ease of scale up
Availability of established practices
equipment for plant harvesting/storage
Elimination of downstream processing
requirement if the plant material
containing the recombinant protein can be
used directly as the protein source
Ability to target protein
production/accumulation to specific plant
tissue
Ability to carry out post-translational
modifications
Disadvantages
Low expression levels often reported
Glycosylation pattern achieved usually
different from that observed on animal
glycoproteins
Lack of industrial experience or data on
large scale downstream processing of plant
tissue
seasonal or geographical nature of plant
growth
presence of toxic substances in plant cell
vacuoles
availability of established alternative
production systems
Modification of Plant Nutritional Content
Amino acid: seed storage proteins contain limited number of amino acids, usually
deficient in one or more amino acids like lysine and methionine. Corn for example
when used as animal feed must be supplemented with soybean meal and or purified
lysine

Lipids: It is possible by genetic engineering to change the degree of unsaturation (no.

of C to C double bonds) and modify canola, to produce modified oil

Use of antisense gene which inhibits the expression of normal gene that leads to the
accumulation of a desired intermediate rather than another product.

Modification of food plant taste and appearance preventing discoloration such as

post-harvest discoloration of fruits and vegetables, therefore control or modify
expression of enzymes which contribute to such discoloration

Sweetness for example in protein monellin, a fruit of an African plant that is 100,000
times sweeter than sucrose and therefore could be a candidate sugar substitute but
which readily loses its sweetness upon heating or when exposed to acid and there are
studies on manipulation of these genes and its expression to address the limiting
characteristics.