Role of the genetic material “A genetic material must carry out two jobs: duplicate itself and control the development of the rest of the cell in a specific way.” -Francis Crick

DNA Deoxyribonucleic acid The information necessary to sustain and perpetuate life is found within a molecule. This is the genetic material that is passed from one generation to the next---a blue print for building living organisms.

History Although we now accept the idea that DNA is responsible for our biological structure, before the mid-1800’s it was unthinkable for the leading Scientists and Philosophers that a chemical molecule could hold enough information to build a human. They believed that plants and animals had been specifically designed by a creator.

History Charles Darwin is famous for challenging this view. In 1859 he published ‘The Origin of Species‘ which expressed that living things appear to be designed, but may actually be the result of natural selection. Darwin showed that living creatures evolve over several generations through a series of small changes.

History In the 1860s Darwin's ideas were finally supported when genetics was discovered by Gregor Mendel. He found that ‘Factors’ determine the characteristics a living thing will express. The genes are passed to later generations, with a child taking genes from both its parents. The great mystery was where and how would this information be stored? 1823-1884 Czech monk

Friedrich Miescher Swiss physician 1844-1895 Search for genetic material: In 1870, a German scientist named Friedrich Miescher had isolated the chemicals found in the nucleus. These were proteins and nucleic acids. (His nuclei came from pus!) Structure was poorly understood Thought that NA is genetic material but further investigation showed that DNA contained only 4 kinds of monomers - seemed like it is too simple for such complicated role. They thought that NA is a structural material Names the substance nuclein because it seemed to come from cell nuclei

History While Miescher found these nucleic acids interesting, and spent a great deal of time studying their chemical composition, he wasn’t alone in believing that proteins were more likely to be the chemicals involved in inheritance, because of their immense variability. Proteins were made up of 20 different building blocks (amino acids), as opposed to the mere 4 building blocks of nucleic acids.

Discovery of a “Transforming Principle” Frederick Griffith, in 1928 - Pneumonia (Diplococcus pneumoniae) infects mice. - Mice develop pneumonia and die. Two types of bacteria: - S bacteria smooth coat – pneumonia - R bacteria rough coat - no pneumonia Coat type is associated with virulence. 1881 - 1941 English army medical officer

History Search for genetic material: 1928 Frederick Griffith: transforming principle

Frederick Griffith’s 1928 Experiment When mouse is injected with dead S strain with intact capsules, the mouse lives – proving that the capsule alone does not kill the mouse. When living non-virulent R strain was mixed with dead virulent cells with intact capsules, the mouse died and S cells were found in mouse. Some substance in the dead S cells transformed the R cells from nonvirulent to virulent. The unknown substance was called the “chemical transforming principle.” Griffith’s Experiment (1920s) • S (smooth) bacterial strain is deadly (virulent) because it has a protective capsule that resists the mouse immune system. • The R (rough) strain lacks the protective capsule, and is destroyed by mouse immune system.

Griffith’s experiment identifying the “transforming principle” Bacterial colonies Rough nonvirulent (strain R) + Rough nonvirulent (strain R) Heat-killed smooth virulent (strain S) Smooth virulent (strain S) Heat-killed smooth virulent (strain S) Injection Results Mouse healthy Mouse dies Mouse dies Mouse healthy Live strain S bacteria in blood sample from dead mouse

Discovery of DNA The extracts of heat-killed S bacteria cells contained protein, RNA and DNA Which of these substances were essential for transformation? How did they figure out which substance was essential for transformation?

KEY PLAYERS Oswald Avery (1877-1955) Microbiologist Avery led the team that showed that DNA is the unit of Inheritance. One Nobel laureate has called the discovery "the historical platform of modern DNA research", and his work inspired Watson and Crick to seek DNA's structure.

Oswald Avery – American bacteriologist 1600s 1800s 1850s 1900s 1950s 2000s 1877-1955 Oswald Avery – American bacteriologist 1943 – proved that DNA carries genes S R DNA Extracted DNA from smooth surface bacteria and introduced it to rough physician and bacteriologist, best known for his discoveries in genetics born in Halifax, Nova Scotia educated at Columbia University's College of Physicians and Surgeons first to show that the agent responsible for transferring genetic information was not a protein, as biochemists of the time believed, but deoxyribonucleic acid (DNA) Avery and his coworkers extracted a substance from a bacterium with a smooth surface and introduced it into a rough-surfaced bacterium. When the rough-surfaced bacteria transformed into the smooth-surfaced type, he knew the substance he had extracted contained the gene that coded for the smooth surface. Avery's team purified this substance and found it was pure DNA. the results were published in 1944, this paper led to more intensive studies of DNA, which eventually revealed it to be the agent of heredity S

Discovery of DNA They decided to use the process of elimination Extracts were treated with either Proteases (to destroy protein) RNase (to destroy RNA) DNase (to destroy DNA) Transformation was due exclusively to DNA They decided to use the process of elimination to figure out which part or parts of the extract were important by destroying each one and seeing what the effect on transformation was. Extracts were treated with either proteases (to destroy protein), RNase (to destroy RNA), or DNase (to destroy DNA). The ability of each of the treated extracts to transform R cells was then tested. Neither the proteases nor the RNase had any effect, but the DNase rendered the extracts inactive - proving that transformation was due exclusively to DNA. Also lipid and protein extraction did not reduce transformation. Oswald Avery published this work in 1943, with Colin MacLeod and Maclyn MacCarthy as co-authors.

Search for genetic material: History Search for genetic material: It wasn’t until 1944 that Oswald Avery and his colleagues, who were studying the bacteria which causes pnuemonia, discovered by process of elimination that bacteria contain nucleic acids, and that DNA is the chemical which carries genes. Despite the conclusive results of Avery’s experiments, the theory of nucleic acids being the genetic material was still not a popular one, but experiments Performed with viruses also showed that nucleic acids were the genetic material and this confirmed Avery’s work.

Alfred Hershey and Martha Chase 1952 1600s 1800s 1850s 1900s 1950s 2000s Alfred Hershey and Martha Chase 1952 used bacteriophage (a virus) to prove that DNA was the hereditary material the bacteriophage was the ideal organism for settling the debate between protein and DNA.

Martha Chase Alfred Hershey

What are viruses? Viruses are organized associations of macromolecules:- nucleic acid contained within a protective shell of protein units . A virus is NOT alive. A virus is NOT made out of a cell.

DNA discovery Hershey-Chase 1952 connecting DNA and heredity. Side by side experiments are performed with separate bacteriophage (virus) cultures in which either the protein capsule is labeled with radioactive sulfur or the DNA core is labeled with radioactive phosphorus. The radioactively labeled phages are allowed to infect bacteria. Agitation in a blender dislodges phage particles from bacterial cells. Centrifugation concentrates cells, separating them from the phage particles left in the supernatant. Results: Radioactive sulfur is found predominantly in the supernatant. Radioactive phosphorus is found predominantly in the cell fraction, from which a new generation of infective phage can be isolated. Conclusion: The active component of the bacteriophage that transmits the infective characteristic is the DNA. There is a clear correlation between DNA and genetic information.

DNA discovery Hershey-Chase 1952

History Search for genetic material: 1952 - Hershey-Chase Experiment

Hershey and

Hershey and

History Search for genetic material: Classic experiments for evidence Griffith: transformation Hershey-Chase: DNA necessary to produce more virus Other supporting evidence DNA volume doubles before cells divide Chargaff: ratio of nucleotides A = T and G = C

Erwin Chargaff – Austrian American biochemist 1600s 1800s 1850s 1900s 1950s 2000s 1929-1992 Erwin Chargaff – Austrian American biochemist # of Adenin units were EQUAL to the # of T G = C

letters, noticing that A Matches to T, and C to KEY PLAYERS Erwin Chargaff (1905-2002) Chargaff discovered the pairing Rules of DNA letters, noticing that A Matches to T, and C to G. He later criticized molecularbiology, the discipline he helped invent, as "the practice of biochemistry without a license", and once described Francis Crick as looking like "a faded racing tout".

James Watson – American ornithologist 1600s 1800s 1850s 1900s 1950s 2000s 1953 James Watson – American ornithologist Francis Crick – British Physicist Complementary double-helical structure of DNA

The Scientists James Watson was an American, born in 1928, was only 24 when the discovery was made. He went to Chicago University at the age of 15. Francis Crick was born in 1916. He went to London University and trained as a physicist. After the war he changed the direction of his research to molecular biology.

Watson went to university in Chicago at age 15, and KEY PLAYERS James Watson (1928- ) Watson went to university in Chicago at age 15, and teamed up with Crick in Cambridge in late 1951. After solving the double helix, he went on to work on viruses and RNA, another genetic information carrier. He also helped launch the human genome project, and is president of Cold Spring Harbor Laboratory in New York.

switched to biology after KEY PLAYERS Francis Crick (1916-2004) Crick trained and worked as a physicist, but switched to biology after the Second World War. After co discovering the structure of DNA, he went on to crack the genetic code that translates DNA into protein. At the time of his death he was studying consciousness at California's Salk Institute.

The Discovery The DNA molecule was discovered in 1951 by Francis Crick, James Watson and Maurice Wilkins using X-ray Diffraction. In Spring 1953, Francis Crick and James Watson, two scientists working at the Cavendish Laboratory in Cambridge, discovered the structure of the DNA a double helix, or inter-locking pair of spirals, joined by pairs of molecules.

How Did They Do That?

The Discovery The seed that generated this was Watson’s presence at a conference in Naples in 1951, where an x-ray diffraction picture from DNA was shown by Maurice Wilkins from King’s College in London. This made a strong impression on Watson – the first indication that genes might have a regular structure.

KEY PLAYERS Linus Pauling (1901-1994) The titan of twentieth-century chemistry, Pauling led the way in working out the structure of big biological molecules, and Watson and Crick saw him as their main competitor. In early 1953, working without the benefit of X-ray pictures, he published a paper suggesting that DNA was a triple helix.

James Watson shared an office with Crick, and the topic of DNA structure naturally arose – particularly how to determine it. They were inclined to follow the methods of Pauling who had designed a helical structure by building a model consistent with the x-ray patterns from fibrous proteins. Like proteins, DNA was built from similar units – the bases adenine(A), thymine(T), guanine(G) and cytosine(C), and so it seemed likely that DNA also had a helical structure. The published x-ray patterns of DNA were not very clear, so contact was made with King’s. Watson attended a DNA colloquium there in November 1951, at which Rosalind Franklin described her results.

Rosalind Franklin- English Chemist 1600s 1800s 1850s 1900s 1950s 2000s 1920 – 1958 Rosalind Franklin- English Chemist the most beautiful X-ray photographs of any substance ever taken (1952) crucial contributions to the solution of the structure of DNA X-ray diffraction images -- double stranded and helical

(1920-1958)

Of the four DNA researchers, only Rosalind Franklin had any degrees in chemistry. She was born into a prominent London banking family, where all the children—girls and boys—were encouraged to develop their individual aptitudes. She attended St. Paul’s Girls School, one of the few schools in London where girls were taught science. Then she proceeded to Newnham College, one of the women’s colleges at Cambridge University. She completed her degree in 1941 in the middle of World War II and undertook graduate work at Cambridge with Ronald Norrish, a future Nobel Prize winner.

She resigned her research scholarship in just one year to contribute to the war effort at the British Coal Utilization Research Association. There she performed fundamental investigations on the properties of coal and graphite. She returned briefly to Cambridge, where she presented a dissertation based on this work and was granted a Ph.D. in physical chemistry. After the war, through a French friend, she gained an appointment at the Laboratoire Centrale des Services Chimiques de l’Etat in Paris, where she was introduced to the technique of X-ray crystallography and rapidly became a respected authority in this field. In 1951 she returned to England to King’s College London, where her charge was to upgrade the X-ray crystallographic laboratory there for work with DNA.

towards the right structure. She went on to do pioneering work KEY PLAYERS Rosalind Franklin (1920-1958) Franklin, trained as a chemist, was expert in deducing the structure of molecules by firing X-rays through them. Her images of DNA – disclosed without her knowledge - put Watson and Crick on the track towards the right structure. She went on to do pioneering work on the structures of viruses.

The Evidence Search for genetic material: James Watson and Francis Crick used this photo with other evidence to describe the structure of DNA. X-ray diffraction photo of DNA Image produced by Rosalind Franklin

In July 1952, Erwin Chargaff visited Watson and Crick and told of his 1947 findings that the ratios of A/T and G/C were statistically equal for a wide variety of DNA’s. Crick became convinced that base pairing was the key to the structure. Prompted by receiving a flawed manuscript on DNA structure from Pauling, Watson again visited King’s and Wilkins showed him a DNA x-ray pattern taken by Franklin showing clear helical characteristics.

OOPS!!! Watson brought back a less-than-accurate account to Cambridge, but Crick produced a three-strand model structure only a week later. Invited to view this,Franklin pointed out that it was inconsistent with her results – it had the phosphate groups on the inside whereas her results showed they were on the outside,and the water content was too low.

Watson & Crick What they deduced from: Franklin’s X-ray data • Double helix • Uniform width of 2 nm • Bases stacked 0.34 nm apart Chargoff’s “rules” • Adenine pairs with thymine • Cytosine pairs with guanine

Watson & Crick What they came up with on their own: • Bases face inward, phosphates and sugars outward • Hydrogen bonding • Hinted at semi-conservative model for replication

History Watson began pursuing the idea of hydrogen bonding by using cardboard cutouts of the four bases. He found that (A+T) and (G+C) could be bonded together to form pairs with very similar shapes. On this basis, a model was built consistent with the Franklin’s symmetry and Chargaff’s results, and Watson & Crick published in April 1953 in Nature accompanied by ones from the Wilkins and Franklin groups. Watson and Crick’s paper ends with the oft-quoted line “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material”.

Watson and Crick with their DNA model

The Nobel Prize Crick, Watson and Wilkins won the Nobel Prize for medicine in 1962. Maurice Wilkins was at King's College, London and was an expert in X-ray photography. His colleague, Rosalind Franklin, did brilliant work developing the technique to photograph a single strand of DNA. She received little recognition for this at the time and died tragically of cancer in 1958, so could not be recognized in the Nobel Award.