杏吧原创

SURPRISINGLY, there is nothing new about biotechnology. The ancient Egyptians
anticipated the penicillin revolution of the 20th century by using mouldy
bread, a good source of antibiotics, as a poultice for infected wounds. And the
earliest evidence for the fermenting of wine comes from chemical analysis of
stains inside an Iranian pottery jar dating back 7000 years to the Neolithic
era.

Biotechnology can be defined as the provision of useful products and services
from biological processes. Often, biotechnology can provide a cleaner, cheaper
and lower-energy way of doing jobs that were previously carried out in chemical
factories, manufacturing or agriculture.

War and yeast

Industry ferments

THE IMPROVEMENTS made in microscope design in the 19th century began to
reveal the role of microorganisms in fermentation, and how they might work for
other human ends beyond brewing. In 1884, Max Delbr眉ck (uncle of the
pioneering molecular biologist of the same name) declared 鈥測east is a
machine鈥� after studying the fungus鈥攁nd so had the first glimpse of
biotechnology鈥檚 potential.

But it was the First World War that really launched biotechnology onto the
industrial stage. Microbiologists began to look beyond the production of ethanol
(an alcohol) from sugar by yeast, and extended the meaning of fermentation to
include all the processes by which microorganisms turn raw materials into
useful products. A process developed in Britain used the bacterium
Clostridium acetobutylicum to turn starchy foods such as rice, potatoes and
maize into acetone and butanol, both good solvents. The alcohol butanol is a raw
material in the production of synthetic rubber, which became a vital resource
for the motor industry after severe shortages of natural rubber during the
pre-war years. Acetone and glycerol (glycerine), which can also be made by
fermentation, are both ingredients of explosives.

The fermentation technology developed then is still important today, although
some of the early products were eventually made more cheaply by the
petrochemicals industry after the 1940s.Then came the discovery of the structure
of DNA by Francis Crick and James Watson in 1953. This led to the development of
genetic engineering by US scientists Paul Berg, Herbert Boyer and Stanley Cohen
in the early 1970s. Genetic engineers manipulate DNA with enzymes to transfer
genes from one species to another. Biotechnologists began to use this new tool
kit to create transgenic organisms of all kinds鈥� microorganisms, plants
and animals. These carry recombinant DNA鈥攃ombinations of genes not
occurring in nature
(Figure 1).

Biotechnology is now a multidisciplinary activity involving chemists,
biologists, engineers and many other specialists. Its scope is enormous.
There are sophisticated new drugs produced in the milk of transgenic sheep,
microbial cocktails that can clean up contaminated land, transgenic plants and
fish that yield more food or resist disease鈥攁nd hundreds of different
microorganisms able to produce fermentation products such as amino acids,
vitamins, enzymes and antibiotics.

All living cells synthesise a wide range of molecules known as metabolites.
Those essential to life鈥攁mino acids (the building blocks of proteins) or
enzymes (proteins which speed up reactions in the body)鈥攁re known as
primary metabolites. Microorganisms and plants can also make nonessential
substances that are called secondary metabolites, such as penicillin and
morphine.

Of the 20 amino acids essential to life, humans can synthesise only 12; the
other eight must be supplied by the diet. Bacteria can synthesise all amino
acids; some of this production can be diverted to human ends. Amino acids such
as lysine and tryptophan are used as human and animal nutritional supplements.
They are also used in the chemical industry鈥攊n the manufacture of the
sweetener aspartame, for example, a dipeptide made from aspartic acid and
phenylalanine.

Microbial enzymes have many uses, from food processing through to equipment
to help doctors make their diagnoses. Enzymes are also the active ingredients of
biological detergents. In the 1960s, Danish scientists discovered that an enzyme
called protease from the bacterium Bacillus licheniformis could break
down protein-based stains, for example, blood or egg. Other enzymes added to
detergents include: amylase dissolves starchy stains from baby food, and lipases
tackle fatty stains including lipstick. Enzymes work best at moderate
temperatures, so the wash is done in cooler water, saving energy.

Some of the most interesting bacteria, from an industrial viewpoint, are the
extremophiles. They get their name because they exist on the edges of
life鈥攗nder tremendous pressure in the deep ocean or in the near-freezing
waters of Antarctica. To survive, extremophiles have evolved interesting
variations on normal biochemistry. For example, Thermus aquaticus,
which was found living at nearly 100 掳C in a bubbling thermal pool at
Yellowstone National Park in the US, is the source of an enzyme called Taq DNA
polymerase. This enzyme is used in a technique called the polymerase chain
reaction (PCR). With PCR, researchers can amplify the quantity of DNA very
rapidly鈥攎aking it possible to analyse tiny amounts of DNA, as in the
forensic analysis of a single hair left at the scene of a crime.

Antibiotics are the main secondary metabolites extracted from microorganisms.
Penicillin, arguably the most important medical discovery of the 20th century,
was first produced in bulk from a strain of Penicillium crysogenum. It
is still produced by fermentation

Species of Alcaligenes bacteria produce carbon storage compounds
such as poly hydroxybutyrate (PHB) which have been made into a range of
biodegradable plastics, known as Biopol. When Biopol is discarded,
microorganisms in the environment quickly break it down to carbon dioxide and
water. Conventional plastics linger unchanged indefinitely in waste tips.

The first stage in a successful fermentation is the selection of a
microorganism which will give a high yield of the desired product. Around 6000
microbial species have been described to date; many thousands remain to be
discovered. Laboratory tests show which species or strain (a variant within a
species) will work well in a given fermentation. Sometimes treating
microorganisms with X-rays or ultraviolet light will product a mutant strain
that gives a very high yield鈥攆or instance, mutant strains give twice as
much of the antibiotic chlortetracycline as the wild types.

Then the microorganisms must be encouraged to deliver peak production.
Although they are not fussy eaters, they often thrive best on a specialised
diet. At the same time, a commercial producer will be looking for a cheap source
of nutrients鈥攊deally a waste product, such as molasses from sugar beet
processing.

An industrial fermenter is just a huge stainless steel tank. It is fitted
with an aerator and a paddle
(or bubbles, Figure 2) to stir the fermenting
mixture鈥攁 鈥渟oup鈥� of working microbial culture and nutrients. Modern
fermenters are also fitted with a control unit that monitors and adjusts
temperature, oxygen level and other conditions.

Temperature control is especially important in fermentation.
Microorganisms generate heat, as do moving paddles. A major risk at all times is the
threat of contamination by the billions of microorganisms in the
surrounding air, on the bodies of the people operating the plant and in the
fermentation ingredients themselves. If a competing microorganism should set up
home in the comfortable environment of the fermenter, it may usurp the working
culture. Nutrients and fermenter alike must be sterilised by pressurised steam
before fermentation begins, and the air must be filtered before it enters the
vessel.

Extracting the product is called downstream processing. If the product
remains stored inside the microbial cells, they must be smashed
open鈥攅ither by a homogeniser (a device like a liquidiser) or by using
detergent to break open the cell walls. Once the product is in the fermentation
broth it may be extracted by an organic solvent鈥攁s penicillin
is鈥攐r by chromatography.

Sometimes it is the process, rather than the product that has commercial
value. Microrganisms are extraordinarily adaptable. Like all living things, they
require a source of carbon. If they live near a source of carbon such as
petroleum waste, which is toxic to humans, they will often use it鈥攂reaking
it down into carbon dioxide and water in the process. We can exploit this:
communities of such toxin-eating microorganisms can be used to clean up
contaminated land sites and polluted waste.

Fungi appear to be good at absorbing toxic heavy metals such as nickel, lead
and even uranium in a process called biosorption. Fungal cell walls contain
chitin, a polysaccharide, that binds tightly to metal ions. Biosorption is used
to concentrate chemical and even nuclear waste, making it easier to handle.
Fungi are made into a mat, through which waste trickles. Heavy metals may be
recovered from filter mats or may remain concentrated in the mats for
disposal.

Productive plants

Lipstick and oil

PLANTS, the planet鈥檚 main food producers, are already a major source of
drugs, flavours and textiles. Although traditionally these useful products have
been extracted directly from leaves and roots, cultures of plant cells, grown
using fermentation-type techniques, may eventually replace agricultural
production.

However, plant cells are not as easy to grow in culture as microorganisms.
The only product to have reached the market so far is the red dye shikonin, from
the root cells of the wild Asian herb Lithospermum erythrorhizon.
Shikonin is used in lipstick, and may also prove to have medicinal
properties.

A more useful application of plant biotechnology is the creation of exact
genetic copies, clones, of an individual with desirable properties. Gardeners
have long known that a whole plant can be created from a single cutting. This is
because plant cells are totipotent, unlike animal cells. Their pattern of gene
expression means that any cell has the potential to become any part of the
plant. Animal cells, with the exception of the fertilised egg or very early
embryonic cells, are pluripotent鈥攖heir daughter cells develop into only a
limited number of cell types.

Making clones from a single part of one plant involves a process known as
tissue culture. The cells are often taken from the meristem, the rapidly growing
tissue at the point of the root or shoot. These are sterilised and placed in a
nutrient solution in a Petri dish where they multiply, forming an
undifferentiated mass of tissue, which is called callus.

If callus is divided up and given plant hormones and nutrients, then roots
and shoots will develop, giving rise to plantlets. These can be grown on in the
usual way鈥攁nd each is genetically identical to the parent. Plant tissue
culture does not require expensive equipment: farmers in Vietnam used it to
clone high-yielding potatoes from the International Potato Centre in Lima, Peru.
Theoretically the method could multiply stocks of endangered and rare plants. On
a larger scale, giant plantations of cloned oil palms are being created in
Malaysia. Palm oil is a source of many products, from plastics to soap. Other
plant oils could be used as chemical feedstock, produced by cloning, to supplant
the world鈥檚 limited supplies of petroleum oil.

Genetic engineering, which transfers genes from one species to another, has
greatly enhanced the power of biotechnology. In principle, genes can be
transferred between any organism鈥攆rom a bacterium to a plant, for example,
or even from humans to other animals鈥攁lthough they may not always be
expressed properly. All the techniques for production and extraction of
products from fermentation can be used in conjunction with genetic engineering.
Most importantly, genetic engineering allows the manufacture of
products鈥攑articularly drugs鈥攖hat could not be obtained in any other
way. And second, it can be used to create organisms, such as plants resistant to
attack by insects, which do not occur in nature.

The first biotechnology product from genetic engineering was human insulin,
used in the treatment of diabetes. Traditionally, insulin was extracted from the
pancreas of pigs or cattle. These insulins differ slightly from the human
version, and some diabetics developed an allergy to them.

Human insulin is made by transferring the human insulin gene into
Escherichia colibacteria. The resulting recombinant bacteria are grown in a
fermenter. The bacteria treat the transferred insulin gene as one of their own
and so, as the bacteria multiply, they manufacture molecules of human
insulin鈥攚hich can be extracted and purified after fermentation. The
products of this kind of fermentation are known as recombinant proteins, or
therapeutic proteins.

Human growth hormone and interferon, a protein used in cancer treatment, are
made in the same way. But bacteria cannot handle the manufacture of the more
complex human proteins, such as haemoglobin or factor 8. These molecules must be
correctly folded into a three-dimensional shape to function. After synthesis in
a human cell, they also acquire several sugar molecules (a process known as
glycosylation). The relatively simple bacterial cell, does not have the
鈥渕achinery鈥� to carry out these tasks.

Recently, nucleated, or eukaryotic, cells have been drafted into the
biotechnology workforce鈥攁nd products such as hepatitis B vaccine and
haemoglobin are already being produced in yeast, while insect and mammalian
cells are increasingly used as hosts. Erythropoietin, which doctors use to
increase the number of red blood cells in patients with very severe anaemia, is
made in cultures of Chinese hamster ovary cells.

Sometimes we can use a whole organism as a 鈥渂ioreactor鈥�, which replaces both
the cell culture and fermenter. Sheep and cattle bearing human genes can produce
therapeutic proteins in their milk鈥攁 process known as pharming. The first
of these products, alpha-1-antitrypsin, is being tested as a treatment for
cystic fibrosis and the lung disease emphysema. Researchers are also
investigating plants as producers of vaccines so that the vaccines may be eaten
in fruit.

One advantage of making therapeutic proteins by genetic engineering is that
it limits the use of potentially infected animal tissue. Many haemophiliacs
became infected with HIV from treatment with factor 8 extracted from
contaminated human blood. This is not a risk when the recombinant version is
used.

Mass market protein

Cure for stroke

GENETIC ENGINEERING also allows the bulk production of proteins that were
previously available only in tiny amounts. One example is interferon, a molecule
produced by the immune system in response to infection. It was once thought that
interferon might be a miracle cure for both cancer and the common cold. Trials
proved disappointing, but a recombinant interferon is now on the market as a
treatment for multiple sclerosis, a condition for which there had been no
therapeutic drugs. Other examples include the clot-busting drugs, such as tissue
plasminogen activator, which save lives when used after heart attacks and,
increasingly, stroke (a condition for which there was no treatment).

Also new is the use of antibodies as drugs. These proteins, produced
naturally by the immune system and manufactured in mammalian cell lines, can
bind to specific molecules in the body and so block their action. They are being
developed for use after transplant surgery to neutralise the action of
substances that would otherwise trigger rejection, and for treatment of septic
shock, cancer and other conditions.

Genetic engineering is also useful in fields other than medicine. Chymosin,
the enzyme used to make cheese, can now be produced by yeast. Recombinant
chymosin can then be used in the production of vegetarian cheese鈥攊n place
of rennet extracted from a calf鈥檚 stomach lining.

Genetic engineering has also been used to create entirely new species of
plants and animals. In plants, the new genes are introduced into cell cultures,
which are then grown up by the methods of tissue culture described above. Many
agriculturally important plant species, including the major food crops such as
soya, rice, wheat and corn, have already been transformed.

In theory, plant genetic engineering could improve the world鈥檚 food supply.
Disease, weeds and predators account for substantial crop losses, however, and
here plant biotechnology can already make an impact. Transgenic plants bearing a
gene for an insecticidal protein from the bacterium Bacillus
thuringiensis are being developed to protect crops from insect attack.
Plants have also been made resistant to a herbicide called Roundup. This means
that a farmer can spray a field of resistant plants knowing that the herbicide
will destroy the weeds, but spare the crop.

More transgenic plants will find their way into our food supply before long.
The only transgenic products in British shops at the moment are tomato paste and
ketchup from transgenic tomatoes. These have been modified so that the gene
responsible for making the fruit go soft has been turned 鈥渙ff鈥�. The
tomatoes stay firm longer and require less energy for processing. It has proved
more difficult to create transgenic animals. Here, a fertilised egg is treated
with the foreign DNA before it is transferred into the womb of a surrogate
mother. Such animals may be an improvement on the original鈥攆ish that grow
much faster or animals with more lean than fatty meat, for example. Researchers
have even created pigs with 鈥渉umanised鈥� organs to avoid rejection. Such
xenografts could make up the shortfall in human donor organs. The animals
bear a human protein called daf (decay accelerating factor) on their organs,
which enables them to evade rejection by the human immune system.

Several strains of transgenic laboratory animals have also been created. With
added human genes, these animals can act as medical models of human disease,
such as sickle cell anaemia and Alzheimer鈥檚 disease, and they may be used to
test new drugs and in basic research.

Biotechnology is an ancient art, but the discovery of genetic engineering has
catapulted it to the forefront of science and industry. Developed safely, it
promises a dramatic improvement in human health and nutrition in the next
century.

* * *

Biosafety concerns

EVEN traditional biotechnology is not without hazard, for microorganisms can
behave unpredictably. In the late 1980s, people taking the amino acid tryptophan
for insomnia and depression began to fall ill, some died. The manufacturers had
changed the microbial strain used in the tryptophan fermentation and,
unknowingly, had selected one that produced a poisonous by-product.

Most safety issues in biotechnology concern genetic engineering. Perhaps the
most controversial area is that of food. Genetically-modified soya and maize
developed in the US are poised to enter the market in Britain. Opponents point
out two safety problems. First, the transgenic plants carry antibiotic
resistance genes. These are inserted early in the genetic engineering process to
select the cells that have taken up the desired 鈥渇oreign鈥� gene. Should these
enter the human gut, they might be passed to gut bacteria, creating a resistant
population and adding to the serious public health problem of antibiotic
resistance. Second, the engineered foods contain 鈥渇oreign鈥� proteins (coded for
by the foreign genes) that might provoke food allergies. The scientific evidence
for either risk appears slim鈥攁nd it is possible to remove the genes for
antibiotic resistance from transgenic plants if they are likely to cause
problems.

When it comes to transgenic plants in the field, we have little experience of
how they might affect the environment when planted on a large scale鈥攖hough
many laboratory experiments and small-scale trials have been done. It is
possible that herbicide resistant genes could be passed on to weeds, enabling
them to grow out of control. And, because herbicides do not affect the
engineered plants, farmers might be less careful in their use of these toxic
substances.