proteins that result from the expression of recombinant DNA within living cells attempted

recombinant proteins recombinant DNA is the general name for a piece of DNA that has been
created by the combination of at least two strands it is possible because DNA molecules from all
organisms share the same chemical structure and differ only in the nucleotide sequence if there is a
protein made then there is a DNA for that protein the protein should be coded by gene this is called
the gene of interest recombinant proteins utilized in the medical world are often referred to as
therapeutic proteins therapeutic proteins provide important therapies for a variety of diseases and
we cast into four groups Group one therapeutic proteins of enzymatic or regulatory activity these
proteins replace a protein that is deficient or abnormal up regulate an existing pathway or provide a
new function or activity group two proteins with special targeting activity these proteins interfere
with the molecule or organism or deliver other molecules Group three proteins as vaccines these
proteins help protect against foreign agents autoimmune diseases and cancer and finally Group four
proteins as Diagnostics these proteins are generally purified and are always recombinant proteins
one of the most important advances in the history of scientific knowledge was the discovery of
DNA by Watson and Crick in 1953 this discovery made possible what is today known as
recombinant technology throughout the 1980s to the 1990s the development of recombinant
proteins was marked by a massive and fatal infection with the HIV virus it was the single cause of
AIDS in the 1980s and hepatitis C virus in the 1990s from 1976 recombinant proteins began being
used in clinical practice beginning with somatic Tosun followed by insulin in 1982 erythropoietin
and M interferon in 1986 tissue plasminogen activator in 1987 and coagulation factors in 1990
today more than 170 recombinant proteins exist in medicine using recombinant DNA technology
enables many proteins to be created outside of the cell recombinant DNA is you to identify map and
sequence genes and determine their function two main ways of creating recombinant DNA is
through molecular cloning which involves replication within living cells and PCR which advise
replication within the test tube through DNA technology recombinant proteins have been used in
many areas they have been using gene therapy which is the introduction of normal genes into cells
in place of missing or defect ones to create genetic disorders drug metabolism recombinant DNA
approaches have recently contributed its role of drug metabolism through heterologous expression
where the enzyme genetic information is expressed into vitro or in vivo through the transfer regime
through antibody and rivet ups plant systems have been recently used for the expression and
development of different antibodies and finally vaccines were comparatively conventional vaccines
have a lower efficacy and specifically than recombinant vaccines to make recombinant proteins the
genes isolated and cloned into an expression vector most recombinant proteins in therapeutic use
are from humans but are expressed in other organisms such as bacteria yeast or animal cells and
culture for this example the gene will be expressed within bacteria so firstly small circular DNA
molecules called plasmids are removed from bacterial cells the plasmids will serve as a vectors in
which they will carry the gene of interest next DNA from the gene of interest is also taken
separately from the cell a restriction enzyme recognizes a specific restriction site of about four to
eight base pairs long on the enzyme it breaks apart the DNA leaving sticky open ends it also cuts
open the plasmids and the gene of interest from the DNA molecule so we now we have three
separate parts to this plasmid the sticky ends of the restriction fragment attached to each other via
base pairing the genes of interests get included into some of the plasmids forming recombinant
plasmids and these are the plasmids that will be transferred into the cells DNA ligase makes the
bonds permanent by attaching nucleotides to each other with phosphodiester bonds it makes the
newly formed DNA and more permanent the plasmids are mixed with bacteria some of them take
up the plasmids in a process called transformation and these are the bacterial cells that we will want
to use the bacteria can be sorted by color isolating which took up the plasmid containing this
specific gene of interest and finally the uncolored bacteria can then be allowed to reproduce
forming a viable amount of pre-teen if the gene for recombinant protein uses codons that are rare in
the host organism that low amount of transfer RNA becomes limited for expression adding the rare
transfer RNA or changing the coding sequence for the gene of interest can resolve this issue the
gene for a commoner protein can also be altered to make the protein more stable increasing the
chance of successful replication changing the n-terminal amino acid or Co expressing a molecular
chaperone or increase protein stability and finally if too much recombinant protein is produced too
quickly it may aggravate inclusion bodies using regulated expression vectors can control the rate at
which recombinant protein is produced the use of recombinant protein technology in medicine has
been very positive overall addressing the many safety issues of the previous methods before this
technology there was a reliance on harvesting proteins from humans or animals to use in treatment
with the very real risks of mistranslations and animal proteins affecting humans in fact human
growth hormone was harvested from cadavers until 1985 resulting in creutzfeldt-jakob disease
being passed on and a significant number of deaths on an even grander scale was a number of
deaths of hemophiliacs from the 1970s once blood clotting factor 8 was able to be extracted
scientifically from blood donors but there's also of course passed along blood-borne viruses like
HIV and hepatitis C resulting in many deaths but from the disaster came an extraordinary
investment in scientific research I guess that led to the identification of HIV as the agent causing
AIDS and the development of a competent factor 8 from an efficiency perspective the development
of PCR has allowed the production of recombinant DNA without the need for vectors or host cells
although not cheap the result is incredibly efficient production of recombinant DNA the science also
continues to evolve with the next stage which is now occurring being third-generation recombinant
proteins which are free from both animal and human proteins these are synthetically produced and
the fourth generation with longer half-lives is in trial investment from both governments and
pharmaceutical companies remains huge in accommodating technology and it's estimated that
medically they will comprise more than 25 percent of the world's pharmaceutical medical markets
by 2025 let's take a look at some of the major drugs using recombinant technology being used at the
moment reading down you can see that insulin is being used for diabetes human growth hormone
for dwarfism and other growth issues interleukin is used in cancer immunotherapy and interferon as
well as for hepatitis A and C and for some forms of leukemia the technology is widely used in
vaccines including influenza hepatitis A and B pertussis diphtheria tetanus and metals it's also used
in for lupus and rheumatoid arthritis for blood cell diseases such as anemia DNA s-1 is used for
cystic fibrosis and coagulation factors very importantly a use for haemophilia a and B you a vaccine
is a weak form of the diseases microorganism that's put into the body once in the body a person
produces antibodies that will protect it in future against the disease the weakened biological form of
the disease in the vaccination is often a surface protein antigen produced by recombinant protein
technology it's worth noting that these are mainly used to protect against secreted toxins rather than
protect against infection by bacteria or virus the vaccinations are based on inactivated toxins to
produce antibodies that neutralize the toxin even though they don't actually kill the infectious
bacteria a yeast called pichia pastoris is the key host system in vaccination recombinant production
due to its high growth rates easy genetic manipulation and its high expression yield examples of
recombinant based vaccinations are hepatitis B diphtheria tetanus whooping cough as well as polio
next we can take a look at the medical replacement of normal proteins or supplements to those that
are in short supply in the body there are many examples of this with the main ones being insulin
human growth hormone factor 8 for blood clotting and FSH insulin though is a great example to
focus in on in diabetes 1 or 2 the body either doesn't produce enough insulin or the cells don't
respond correctly to it in either case the body needs more insulin than it's produced to ensure that
glucose doesn't build up in the blood and that the cells get an adequate energy source recombinant
insulin was the first approved genetically engineered drug for use because the insulin gene is at the
top of the chromosome it was very easy to isolate and by 1982 it was produced in bacteria since the
exact order of insulins amino acids are known an amino acid sequencing machine connects the
amino acids together in the correct order and synthesizes the DNA it's then put into a host usually
that's Ecole a or yeast to grow and then separated and purified this can be done on a very large scale
resulting in a very efficient high production product that's relatively cheap to produce as I'm sure
you know there are an estimated 1.2 million people aged 2 years and over which is 5% of the
population in Australia diagnosed
 with diabetes according to the Australian Department of Health it gives you an idea just how
important this technology is for so many people the next exciting stage of recombinant insulin is
analog insulin this is where the amino acid sequence is changed or tweaked slightly but not too
much so that the still cell still accepts its insulin the chemical nature changes so that it actually
repels other land with cancer a primary example is cytokines that have been manufactured using
recombinant technology these are then injected so that they become present in a much greater
number in the body than would naturally be found cytokines remember other proteins involved in
the immune system which carry signals between cells and alter their behavior in many ways
specifically they control the body's cohesive response to pathogens bacteria and to toxins one way
to tackle some forms of cancer and leukemia is by immunotherapy which is a term for treatments
that don't directly target the cancer cells themselves but rather stimulate the immune system to fight
the cancer two good examples of these recombinant cytokines are interferon type 2 which is known
as AI fn2 and interleukin 2 which is called aisle 2 if we take a look at afn 2 firstly it's used to
increase the number of immune cells actually produced this stimulates an increase in the number of
both macrophages and dendritic cells to attack any unhealthy cancer cells it's used as a treatment in
specific diseases such as kidney and bladder cancers the circulatory cancers like leukemia and
lymphoma and also for melanomas il-2 however is different in that it stimulates increased
lymphocyte production of beta cells and T cells to fight cancer beta cells remember make antibodies
and the increased number of antibodies can then bind an increase destruction of any abnormal cells
killer T cells however can destroy abnormal cancer cells in two ways by both cell self-destruction
which is called apoptosis or by releasing toxic chemicals although both of these are reasonably new
technologies the results are so far very promising so as I hope you can see recombinant proteins
have led to massive innovations and medical treatment options and already save many many lives hi
I'm Ella Carrick and I'm going to be talking about the future recommendations of recombinant
proteins to start off with it I'm going to talk about the shortcomings of this technique some of the
shortcomings of the use of recombinant proteins in medicine include ethical concerns using these
proteins can be seen as going against nature religious beliefs and safety there are concerns whether
changing the genetic material of human beings is ethical and if it's justifiable also whether it's okay
to select good genes such as appearance intelligence or abilities what would be considered to be
normal if such genes for chosen would those without these selected good genes be discriminated
against this causes some social concerns with many people naming designer babies this might mean
that those people that are wealthy or higher socioeconomic would be able to afford to selecting
these genes or the preferred genes and those are less fortunate would not be able to in some case it's
seen as unethical to go against particular religious beliefs and values as we are placing the God the
role of God which can portray the image of God as flaw it can also be potentially dangerous for
individuals when changing natural DNA sequences including mutations and reducing productivity
or proteins which will be discussed in more detail from the time of discovery since the 1950s when
selfies from a monkey to successfully generate a vaccine technology as vans and new techniques
are available today synthetic genomes are capable of self-replication intricate bacterial cell
genetically engineered cells themselves can be used to treat patients that have suffered a stroke for
example there are also advances in 3d printing technology which is used for skin printing used for
major burns and wounds and it's been highly successful also synthetic human insulin is also being
produced as well as sequencing of human whole genomes the most recent development is the CRI
SPR which is recombinant DNA technology that is used to target destruction of genes in human
cells genetic engineering may assist to overcome many threats to the human life such as disease
malnutrition to delimited food environmental issues caused by organization in the industrialization
however there are some issues that arise from this technique the production of recombinant
pharmaceuticals encounter arranger obstacles which restrict them from producing functional
proteins some of these problems that can arise include cell stress response post translational
modifications low solubility instability of paralytic activities and resistance in expressing new genes
mutations can also occur when the DNA is maintenance processes fail resulting in permanent deep
change in the DNA mutations that occur in humans at a genetic level reduced productivity of
proteins the mutation can adjust stroy and organism if it occurs on a vital position on the DNA
sequence these issues need to be overcome by specific gene enhancement according to the
individuals genome the amino acid sequence of a recombinant human protein express an e coli may
be identical to the human wild-type and yet the protein may lack or have a decrease biological
activity bacteria being example very chaotic organisms differ from the higher organisms you know
they are not able to perform a majority of protein post translational modifications which the cell
would require specialized cell compartments potential future applications are extensions of this
technique the latest advances in protein engineering technologies have allowed drug developers and
manufacturers to fine-tune and exploit desirable functional cara sticks of proteins of interest while
maintaining product safety and efficiency as free competent proteins have been approved to treat
patients with cancer autoimmune disease disorders and genetic disorders it has already improved
the health outcomes for many individuals further advances of vaccines and pharmaceuticals using
recombinant proteins for such disease and disorders will continue to improve health and potentially
prevent death in some patients researchers are confident to report that recombinant proteins will
continue to develop and assess the critical diseases and illnesses that put human lives at risk further
studies and technology will continue to improve techniques and make it more accessible for your
patients however extensive research into recombinant proteins is required to overcome issues and
concerns for humans in their environment education and awareness needs to be delivered to the
population to offer understanding about recombinant proteins while respecting people's views and
values the following two slides will contain the references used for this presentation thank you