DNA: Discovery of Genetic Material

• James Watson and Francis Crick discovered DNA structure in 1953, building on prior work.

Discovery of the Genetic Material

• In 1928, Frederick Griffith's experiment suggested DNA as the genetic material.
• Griffith studied Streptococcus pneumoniae strains: smooth (S) and rough (R).
• S strain has a sugar coat, forms smooth colonies, and causes pneumonia.
• R strain lacks a sugar coat, forms rough colonies, and does not cause pneumonia.
• Injecting live S strain into a mouse kills it, while live R strain does not.
• Heat-killed S strain is not lethal, but heat-killed S mixed with live R strain kills the mouse.
• Griffith concluded that the R strain transformed into a live S strain.
• In 1931, Oswald Avery determined that DNA transformed the R strain into S strain.

DNA as Genetic Material

• Despite Avery's findings, many scientists doubted DNA as the genetic material.
• In 1952, Alfred Hershey and Martha Chase proved DNA was the genetic material.
• Hershey and Chase used bacteriophages, viruses infecting bacteria, made of DNA and protein.
• They labeled bacteriophage DNA with radioactive phosphorus (32P) and protein with radioactive sulfur (35S).
• Bacteriophages injected their genetic material into bacteria and were then separated.
• Only the labeled DNA entered the bacterial cells, confirming DNA as the genetic material.

Discovery of DNA Structure

• After the Hershey-Chase experiment, how nucleotides arranged or DNA followed Chargaff's rule remained unknown.
• Rosalind Franklin, Maurice Wilkins, Francis Crick, and James Watson solved the DNA structure.
• Franklin's X-ray diffraction image, Photo 51, showed DNA as a double helix.
• Watson and Crick used Franklin's data to build a DNA model with deoxyribose and phosphate strands.
• Bases inside the molecule pair: cytosine with guanine, thymine with adenine.
• In 1953, Watson and Crick published the DNA structure in Nature, supported separately by Wilkins and Franklin.

Nucleotides

• DNA is made of nucleotides, each with deoxyribose sugar, a phosphate group, and a nitrogenous base.
• The four nitrogenous bases in DNA are adenine, guanine, cytosine, and thymine.
• RNA is also made of nucleotides, but contains ribose sugar, a phosphate group, and the nitrogenous bases adenine, guanine, cytosine, and uracil.
• Adenine and guanine are double-ringed purine bases.
• Thymine, cytosine, and uracil are single-ringed pyrimidine bases.
• Erwin Chargaff found that guanine nearly equals cytosine, and adenine nearly equals thymine (Chargaff's rule: C = G, T = A)

Structure of DNA

• DNA resembles a twisted ladder with deoxyribose and phosphates as the rails.
• Base pairs (cytosine-guanine or thymine-adenine) form the rungs.
• Purine binds to pyrimidine to give constant rung width.
• DNA strands run in opposite directions, labeled 5' to 3' and 3' to 5'.

Semiconservative Replication

• Every time a cell divides, it must copy its DNA.
• Semiconservative replication involves two DNA strands separating and serving as templates to make DNA molecules with one parental and one new strand.
• DNA replication happens in interphase of mitosis and meiosis, with three phases: unwinding, base pairing, and joining.
• DNA helicase unwinds the double helix in the first phase.
• Single-stranded binding proteins hold the strands apart.
• RNA primase adds an RNA primer to each DNA strand.
• DNA polymerase adds nucleotides, bonding them to the parent strand and forming base pairs in the next step.

DNA Strands

• One DNA strand runs 3' to 5', and one runs 5' to 3'.
• The leading strand is continuously made longer as the original DNA unwinds.
• The lagging strand grows through Okazaki fragments in the 3' to 5' direction.
• DNA polymerase removes RNA primers, replacing them with DNA nucleotides.
• DNA ligase connects DNA nucleotides.

DNA Replication

• Eukaryotic DNA replication occurs simultaneously at many sites, appearing as bubbles in the DNA.
• Prokaryotic DNA is circular and replication starts at one point, proceeding bidirectionally until the entire strand is copied.

Central Dogma

• Proteins act as cell building blocks and enzymes, with instructions for their creation found in DNA.
• Information route: DNA to RNA to proteins forms the central dogma of biology.

Types of RNA

• RNA, like DNA, is a nucleic acid but contains ribose sugar, uracil replaces thymine, and usually exists as a single strand.
• Messenger RNA (mRNA) carries instructions for protein synthesis from the nucleus to the ribosome.
• Ribosomal RNA (rRNA) forms part of the ribosome in the cytoplasm.
• Transfer RNA (tRNA) transports amino acids from the cytoplasm to the ribosomes.

Transcription

• During transcription, mRNA is made from DNA.
• DNA unzips in the nucleus, and RNA polymerase binds, producing mRNA from 5' to 3'.
• Uracil replaces thymine, forming a complementary mRNA strand that then moves out via nuclear pores.

RNA Processing

• Pre-mRNA contains all DNA code, needing processing before leaving the nucleus.
• Introns (interrupting sequences) are cut out.
• Exons (protein-coding sequences) remain.
• A protective cap is added to the 5' end and a poly-A tail (string of adenines) to the 3' end.

The Code

• The sequence of bases determines the way DNA varies among organisms.
• DNA must provide codes for at least 20 amino acids.
• A three-base code called a codon transcribes into the mRNA code.
• Three codons (UAA, UAG, UGA) are 'stop' codons, ending transcription.
• AUG codes for methionine and acts as the start codon.

Translation

• mRNA leaves the nucleus, connects to the ribosome, where it is translated into proteins.
• tRNA molecules act as 'interpreters'.
• Each tRNA folds like a cloverleaf, binds to a specific amino acid at the 3' end, and carries an anticodon.
• Anticodon is complementary to mRNA codon and reads from 3' to 5'.
• The ribosome, composed of large and small subunits, facilitates this process.

Protein Production

• A tRNA with anticodon CAU carrying methionine binds to the start codon AUG.
• A tRNA complementary to the mRNA codon binds to the P site, followed by introduction of a second tRNA to the A site.
• The ribosome joins amino acids, releasing the tRNA to the E site to exit.
• Ribosome shifts along mRNA; tRNA in the A site moves to the P site and continues adding/joining amino acids.
• The process stops when the ribosome reaches a stop codon which releases mRNA.

One Gene-One Enzyme

• Each gene codes for one enzyme "one gene–one enzyme" hypothesis.
• Beadle and Tatum's studies of Neurospora confirmed that a gene can code for an enzyme
• Some enzymes contain polypeptide chains, shifting the hypothesis to: each gene codes for one polypeptide.

Prokaryote Gene Regulation

• Cells use gene regulation in response to the environment.
• Prokaryotes utilize operons: DNA sections with genes for specific metabolic pathways.
• An operon contains an operator (on/off switch), a promoter (RNA polymerase binding site), and a regulatory gene.
• The tryptophan (trp) operon is a repressible operon.
• Low tryptophan levels allow RNA polymerase to bind to the operator, activating transcription of five genes that make tryptophan.

Trp Operon Regulation

• High tryptophan levels bind to the repressor protein; keeping RNA polymerase from binding and turning off.
• The lac operon is an inducible operon with genes coding for lactose digestion enzymes.
• An inducer (like lactose) binds to and inactivates the lac repressor, enabling RNA polymerase binding.
• If lactose is unavailable, the repressor protein binds to the operator to block transcription.

Eukaryote Gene Regulation

• Eukaryotes use complex gene control methods since they have more genes.

Transcriptional Control

• Transcription factors controls gene activation and protein production.
• Transcription factors bind RNA polymerase to the promoter.
• Other transcription factors control the rate of transcription.

Hox Genes

• Homeobox (Hox) genes encode transcription factors and control differentiation.
• Hox genes develop different body parts during embryo development

RNA Interference

• Eukaryotic genes regulate with RNA interference (RNAi).
• Interfering RNA molecules bind to protein complexes that breaks down one strand of RNA.
• The resulting RNA and protein complex prevents mRNA translation.

Mutations

• A change in the cell's DNA sequence results in a mutation.
• Mutations change proteins.
• Point mutations happen when a single nucleotide changes.
• Substitution: one base is exchanged for another.
• Missense mutation changes the DNA code to code for the wrong amino acid.
• Nonsense mutation changes an amino acid codon into a stop codon, shortening the protein, as seen in muscular dystrophy.

Insertion/Deletion Mutation

• Insertions and deletions: nucleotides in DNA are either added or lost causing a frameshift mutation.
• Frameshift Mutation: the ribosome misreads the codons.
• Frameshift mutations cause cystic fibrosis and Crohn's disease.
• Large DNA mutations delete/move pieces of a chromosome.
• Tandem repeats increase in number (Fragile X syndrome, Huntington's disease).
• Small mutations may alter how proteins fold, causing functional changes and resulting in genetic disorders.

Causes of Mutation

• Some mutations occur during DNA replication when DNA polymerase makes a mistake.
• Mutagens cause other mutations by damaging DNA (chemicals, radiation).
• Thymine bases bind together due to UV radiation, creating a kink and preventing replication.

Inheritance of Mutations

• Mutations in somatic or body cells are not passed on.
• Mutations in sex cells pass to offspring.
• Mutations leads to no change, while others cause disease.