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DNA, RNA, AND PROTEIN SYNTHESIS Chapter 10 IMPORTANCE OF DNA Why is DNA important? • DNA contains the instructions for how the cells of all living things construct proteins, including enzymes. • We have discussed how proteins and enzymes are important to living things. • Your skin, muscles, and bones are made of proteins. • Any chemical reaction in your body requires enzymes, which are proteins. • DNA in universal • Me, you, grass, ant, mushroom, pond scum…all based on DNA – sugar, phosphate, bases…the bases are just in different orders! Why do we study DNA? • It is the central importance to all living things on Earth! • It is the answer to all those questions we ask…why do I look like my parents, how do I get sick, will I get cancer, why do I act the way I do??? • Medical benefits/biotech – cures for diseases, understanding cancer and birth defects, genetic testing, production of desired product (i.e. Insulin) • Better food crops (?) GMO’s are genetically modified organisms (usually food); taste better, grow larger • CSI – DNA provides the evidence to prove a criminal’s involvement in a crime Genetic Engineering • As genetics allows us to turn the tide on human disease, it's also granting the power to engineer desirable traits into humans. What limits should we create as this technology develops? • Genetics is our next unit… HISTORY OF DNA History of DNA • There were many experiments conducted early in the 20th century that provided evidence to support that DNA, not proteins or RNA, is the genetic material in all living cells. • 1928 – Frederick Griffith • 1944 – Avery, McCarthy, & MacLeod • 1950 – Erwin Chargaff • 1952 – Hershey & Chase • 1953 – Franklin & Wilkins – Watson & Crick History of DNA • 1950 – Erwin Chargaff • Proposes two rules regarding composition of DNA: 1. Number of complimentary bases is always equal in DNA • #Cytosine always equal #Guanine • #Adenine always equal #Thymine • In human DNA, bases are always found in the following percentages: A=30.9% and T=29.4%; G=19.9% and C=19.8% • This data suggested the base pairing among DNA (although Chargaff never stated it! He did tell Watson and Crick though!) 2. Composition of DNA varies from one species to another in the relative amounts of A, G, T, and C bases. This evidence of molecular diversity, which had been presumed absent from DNA, made DNA a more credible candidate for the genetic material than protein. Chargaff’s Rules Conclusion • By 1952, the experiments of Griffith, Avery and his colleagues, and Hershey and Chase led most scientists to accept (based on the evidence we’ve discussed) that DNA was the hereditary material of living things. • Chargaff also shed light on the composition of DNA. • It was yet to be determined… • The structure of DNA… • How is replicates… • How it has the ability to store and transmit information in a cell… History of DNA • 1953 – James Watson, Francis Crick, Rosalind Franklin, & Maurice Wilkins • Franklin & Wilkins discover DNA is helical by use of X-ray diffraction • “Photo 51” • Watson and Crick obtain the famous photograph without permission • They are able to build a model of DNA with help of photo and other information • They ALL published their findings in the same issue of Nature. 1962 Nobel Prize winners – Physiology & Medicine • Rosalind Franklin died of ovarian cancer in 1958 at the age of 37. • Sadly, Nobel Prizes are only awarded to the living. STRUCTURE OF DNA DNA: DEOXYRIBONUCLEIC ACID Recall 4 classes of macromolecules 1 class are Nucleic Acids Examples are DNA & RNA Monomers are small units that make up larger units called polymers. Nucleic acids are polymers. This means DNA is a polymer. Who can remember what the monomer of a nucleic acid is? DNA Structure • NUCLEOTIDES! • Nucleotides have 3 parts: 1. Nitrogenous base (there are 4 kinds) 2. Phosphate Group 3. 5 carbon sugar: Deoxyribose DNA Structure • Nitrogenous bases: • Contain nitrogen and is a base • Purines – double ringed • Adenine (A) • Guanine (G) • Pyrimidines – single ringed • Cytosine (C) • Thymine (T) Hydrogen Bonding • A purine (A or G) will easily form a hydrogen bond with a pyrimidine (T or C). This is because of their structure and the availability of bonding sites. • Hydrogen bonds are weak but there are millions and millions of them in a single molecule of DNA. phosphate One strand of DNA is a long polymer of nucleotides. One strand of DNA has many millions of nucleotides! nucleotide deoxyribose bases DNA is Double-Stranded • DNA is made up of 2 straight chains of nucleotides • The bases on each of those chains are attracted to each other and form hydrogen bonds • The force of thousands or millions of hydrogen bonds keeps the two strands of DNA held tightly together DNA is a Double Helix • Double helix means two sides twisted in a spiral. • Backbone – alternating sugars and phosphates. • Bases down the middle held together by hydrogen bonds DNA Structure • If DNA was a spiral staircase… • The alternating sugar and phosphates would be the hand rails. • The bases would be the steps • But, they would be weak steps as they are only held together by hydrogen bonds Base Pairing Rules Adenine ALWAYS pairs with Thymine Cytosine ALWAYS pairs with Guanine • Each pair contains one single and one double ringed base • They “compliment” each other Complimentary Base Pairing DNA Structure • Because of the base pairing rules, one strand of DNA is complementary to the other strand (otherwise they would not stick together!) • The order of the nitrogenous bases on DNA is called its base sequence. • So if one strand has a base sequence of TGCC, the other strand will have ACGG. Let’s Practice • Write the complimentary strand for… TGACCGAT ACTGGCTA TGGCCAATATA ACCGGTTATAT DNA REPLICATION DNA Replication • DNA Replication is the process by which DNA is copied during interphase of mitosis before the cell divides. DNA Replication • First, enzymes called Helicases separate (or “unzip”) the two strands of DNA • Helicases break hydrogen bonds • The Y-shaped region formed as bonds are broken is called the replication fork DNA Replication • Next, enzymes called DNA polymerases add complimentary nucleotides to the separated strands of DNA • Nucleotides are found floating freely in the nucleus • The addition of new nucleotides occurs in opposite directions on each strand (one toward the replication fork and one away from it) DNA Replication • In Eukaryotes, several replication forks form on the DNA and the process continues until all of the DNA has been replicated. • If only 1 was formed it would take too long to replicate DNA (53 days for humans!) • When replication is finished, there are 2 DNA molecules, each has one old strand and one new strand • This is called semi-conservative DNA Replication • Replication is usually very accurate • There is only about 1 error for every BILLION nucleotides added! • The reason is that DNA Polymerases also “proofread” the DNA and fix any errors during replication DNA Replication • If an error does occur, it results in a different nucleotide sequence in the new DNA strands • This is called a mutation • A change in even one nucleotide COULD be very harmful to an organism (for reasons we will see later) • Some mutations can affect the growth of cells, causing growth to accelerate, this results in cancer • Changes can be good! Mutations sometimes create new variations that may be advantageous and lead to evolution. UNDERSTANDING THE GENETIC CODE PROTEIN SYNTHESIS Protein Synthesis • DNA is the “code” for hereditary characteristics. • The genetic code is how organisms store hereditary information and translate it into amino acids. Protein Synthesis • DNA codes for all of the bodies proteins • Genes are sequences located in the DNA that code for specific characteristics. • For example, the code (or gene) for the production of the protein melanin is in your DNA and creates your hair and skin color. • The code or recipe for all of the enzymes that help you digest your food is located in your DNA. The Human Genome • A genome is the complete genetic content of an organism. • HGP was an international research project with the goal of determining the sequence of our base pairs in DNA • Started in October 1990 and completed in June 2000! • Biologists have deciphered 3.2 billion base pairs in the 23 human chromosomes • These sequences code for about 22,000 genes • We are still researching which sequences code for which genes. Protein Synthesis • The “code” or “recipe” within DNA cannot be read directly… • DNA cannot leave the nucleus and proteins are made by ribosomes in the cytoplasm of cells • So the code is transcribed (copied) and translated (turned into something useful) by ribonucleic acid (RNA) Protein Synthesis • Remember, proteins make us who we are • They are responsible for cell structures, chemical reactions occurring in us (enzymes), and for the hereditary characteristics that we have (such as eye color) • The building blocks (or monomers) of proteins are amino acids • DNA holds the recipe for the amino acid sequence of all the proteins we need to make RNA & Protein Synthesis • RNA stands for: Ribonucleic acid • RNA directs protein synthesis, which is the making of proteins from DNA Comparing DNA & RNA • Both are made of nucleotides • Both are needed for protein synthesis Comparing DNA vs RNA DNA RNA • Double-stranded • Single-stranded • Sugar: Deoxyribose • Sugar: Ribose • Bases: A, T, C, G • Bases: A, U, C, G • Thymine • URACIL replaces Thymine • Much shorter than DNA TYPES OF RNA There are 3 types of RNA involved in protein synthesis… Messenger RNA • Abbreviated: mRNA • Carries the genetic instructions from the DNA to the ribosomes Ribosomal RNA • Abbreviated: rRNA • Part of the ribosome • Remember ribosomes make proteins Transfer RNA • Abbreviated: tRNA • Transfers the amino acids to the ribosomes to make proteins January 8, 2013 Core Objective • How are DNA and RNA different? • Agenda: Notes, RNA coloring • Homework: RG, Prelab Catalyst PROTEIN SYNTHESIS If the instructions for making proteins are housed in the nucleus in DNA, then there must be a way for the instructions to get to the ribosomes where proteins are made… PROTEIN SYNTHESIS: TRANSCRIPTION Step 1: Transcription (to re-write) DNA is too large to go from the nucleus to the cytoplasm, so only pieces of DNA are copied into RNA. This RNA then travels from the nucleus to the cytoplasm. Transcription • Step 1: • An enzyme called RNA polymerase binds to a genes promoter region • A promoter is just a specific nucleotide sequence where the RNA polymerase can attach • DNA begins to uncoil Transcription • Step 2: • The RNA polymerase adds complimentary nucleotides resulting in a straight chain RNA molecule • The DNA code determines what bases will be added •A → U T → A G ↔ C • For example: ATCCGTT • RNA: UAGGCAA • *Remember, RNA does not have thymine, it has Uracil!! • DNA: Transcription • Step 3: • Copying of DNA continues until the RNA polymerase reaches a termination signal • a specific sequence of nucleotides that tells the RNA polymerase to “STOP” and release the RNA and DNA • The RNA is mRNA, because it is the messenger of the “code” from the DNA to the ribosome Transcription - Overview RNA polymerase – starts unwinding the DNA when it attached to the “promoter” (specific nucleotide sequence) 2. RNA polymerase – adds RNA nucleotides 3. RNA polymerase – stops when “termination signal” (specific nucleotide sequence) is reached. 1. Last slide – not in note outline Before going on… let’s review • Transcribe this DNA strand into mRNA: ATTGGCTGCTTAGC UAACCGACGAAUCG PROTEIN SYNTHESIS: TRANSLATION Step 2: Translation (to make useful) The RNA is then made into something useful, like assembling amino acids into proteins in the ribosome. Translation • Once the newly made mRNA leaves the nucleus it attaches to a ribosome at the promoter region. • Ribosomes will “read” 3 nucleotides in the mRNA code at a time • These 3 nucleotides are called codons (or triplets) • Each codon codes for either an amino acid, a START signal, or a STOP signal Translation • For example, the sequence AUG codes for the amino acid Methionine and means START (it is the only one that means start) • ALL mRNA molecules start with AUG, otherwise, they would not have a start region for protein synthesis Translation • So, in translation, the mRNA is translated into amino acids, which are put together to form proteins (or polypeptides) • The translation occurs with the help of tRNA, which carries the amino acids Translation • When the ribosome reads the start sequence (AUG), a tRNA molecule comes along with the anticodon • The anticodon is the complementary sequence, which would be UAC. • The complementary bases bond with each other and the amino acid methionine begins the protein synthesis within the ribosome • tRNA transfers amino acids to the ribosome Translation • There are only 20 amino acids • Most amino acids have more than one codon • For example, Leucine’s codons are UUA, UUG, CUU, CUC, CUA, and CUG • But each codon codes for ONLY 1 amino acid • For example, CUU only codes for Leucine and nothing else mRNA Codon Chart Translation • After the start sequence, the ribosome moves to the next codon. • Let’s say the next codon is GUC • Now a tRNA that has the anticodon CAG would attach to the ribosome and it would carry the amino acid Valine. • The amino acid Valine would attach to the Methionine from before (now we have a dipeptide!) Translation • This process continues and the polypeptide grows until the STOP codon is reached • UAA, UAG, and UGA are the only stop codons • The protein, ribosome and all RNA is released to perform other needed functions Protein Synthesis - Overview • Amino Acids are listed by their CODON!!! • That would be the 3 nucleotide sequence on the mRNA Protein Synthesis - Overview •This is an mRNA strand •Figure out what the DNA code was that it came from Protein Synthesis - Overview A→T U→A C↔G mRNA Strand: AUG/ACG/GAG/CUU/CGG/AGC/UAG DNA Strand: TAC/TGC/CTC/GAA/GCC/TCG/ATC Protein Synthesis - Overview •Now figure out the anticodons (which will be found on the tRNA) Protein Synthesis - Overview A→U U→A C↔G mRNA Strand: AUG/ACG/GAG/CUU/CGG/AGC/UAG tRNA Strand: UAC/UGC/CUC/GAA/GCC/UCG/AUC Protein Synthesis - Overview •1 - UAC •2 - UGC •3 - CUC •4 - GAA •5 - GCC •6 - UCG •7 - AUC Protein Synthesis - Overview • Now use the CODON chart to figure out the amino acid sequence • Remember to use the codons from the mRNA to determine amino acid sequence. Protein Synthesis - Overview •1 – Methionine (start) •2 - Threonine •3 – Glutamic Acid •4 - Leucine •5 - Arginine •6 - Serine •7 - STOP Let’s Break the Genetic Code 1. Start with DNA: TA C TA G C TAA C C 2. Write the complimentary strand for mRNA AUGAUCGAUUGG 3. Identify the codons on the mRNA AUG-AUC-GAU-UGG 4. Identify the anticodons on the tRNA UAC-UAG-CUA-ACC 5. Identify the amino acid sequence from the mRNA Met - Iso - Asp - stop January 9, 2013 Core Objective • How are DNA and RNA different? • Agenda: Notes, RNA coloring • Homework: RG, Prelab Catalyst MUTATIONS Genetic Mutations • Any change in the DNA sequence is called a mutation. • Mutations can effect body cells • Example = CANCER • Mutations can effect reproductive cells • These are called germline mutations and can be passed from parent to child. • If a child inherits a germline mutation from their parents, every cell in their body will have this error in their DNA. • Germline mutations are what cause diseases to run in families Not All Mutations are BAD! • Some mutations result in characteristics that give the organism a greater chance of survival. • Example: Sickle cell anemia (deflated look of red blood cells) is caused by a mutation, however it is beneficial to people in Africa who often contract malaria – the parasite can no longer attach to their red blood cells, therefore they aren't affected Genetic Mutations • Types of DNA Mutations: • Point • Frameshift • Inversion Point Mutation • A change in a single base pair in DNA • Silent Mutation – changes a base pair, but no change to amino acid sequence • Example: CUU ( Leucine ) to CUC ( Leucine ) • Missense Mutation – changes a base pair, therefore changing amino acids • Example: AGU (Serine) to AGA (Arginine) • Nonsense Mutation – changes a sequence to a stop codon. • Example: AGA (Arginine) to UGA (Stop) Frameshift Mutation • A single base pair in DNA is deleted or added. • Every codon after the deleted or added base would be different. • This type of mutation can make the DNA meaningless and often results in a shortened protein. Inversion • In an inversion mutation, an entire section of DNA is reversed. • A small inversion may involve only a few bases within a gene, while longer inversions involve large regions of a chromosome containing several genes. Examples • Example: • THE DOG BIT THE CAT. • THE DOB ITT HEC AT. • THE DOG BIT THE CAT. • THE DOG BIT THE CAR. • Frameshift • Point • The fat cat ate the wee rat. • The fat tar eew eht eta tac. • Inversion Causes of Mutations • Spontaneous: • Mistake in base pairing during DNA replication • Mutagen – agent that causes DNA change • High energy radiation • X rays • Chemicals • Dioxins, asbestos, benzene, cyanide, formaldehyde • High temperatures THE END Some Great Resources • DNA Replication animations: • http://207.207.4.198/pub/flash/24/menu.swf • http://www.ncc.gmu.edu/dna/repanim.htm • Protein Synthesis animations: • http://www.lewport.wnyric.org/JWANAMAKER/animations/Prot ein%20Synthesis%20-%20long.html • http://www.wisconline.com/objects/index_tj.asp?objID=AP1302 Some Great Resources • http://nobelprize.org/educational_games/medicine/dna _double_helix/index.html • DNA games