Genetics 3000 Exam 4 Study Guide
Genetics 3000 Exam 4 Study Guide 85033 - GEN 3000 - 002
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85033 - GEN 3000 - 002
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This 12 page Study Guide was uploaded by Lisa Blackburn on Sunday April 3, 2016. The Study Guide belongs to 85033 - GEN 3000 - 002 at Clemson University taught by Kate Leanne Willingha Tsai in Fall 2015. Since its upload, it has received 197 views. For similar materials see Fundamental Genetics in Biomedical Sciences at Clemson University.
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Date Created: 04/03/16
Exam 4 Study Guide (Chapters 1315) Chapter 13: Translation and Proteins I. Ribosomes: a. Machines of protein synthesis. Consist of protein and RNA b. rRNA genes: make complexes with proteins, creates large and small subunits to make a fully attached “ready to go” ribosome. c. Prokaryotes and Eukaryotes both have small and large subunits i. Eukaryotic subunits are larger in general than prokaryotes ii. Have to have both subunits to be able to assemble a ribosome which is needed for translation to take place II. Translation: divided into stages a. Charging: the binding of the amino acids to tRNA charges the ribosome i. Once tRNA is charged, it get used and loses its charge. It then can be recharged and reused again ii. Happen continuously throughout translation b. Initiation: what starts the whole process of translation, getting the ribosome into the right location on the mRNA c. Elongation: hooking together all of the amino acids d. Termination: stopping the process III. tRNA Charging: a. There is a tRNA for each amino acid i. NOTE: because of the wobble position, one tRNA picks up on several codons, resulting in seeing fewer than what would be calculated as possible b. Modification that tRNA can undergo: can create a modified base, creating a specialized base that will allow for it to recognize a C or a G base i. Helps with the wobble c. All of these tRNAs have the same attachment site on top that says that it needs an amino acid i. Amino acids cannot tell which tRNA is the correct one for them to bind to ii. Will use a key enzyme to direct the right amino acid to the right tRNA 1. Key enzyme: aminoacyltRNA synthetases****************** 2. Different one for each amino acid, finds all of the tRNA that binds to a certain amino acid, and then will connect them 3. tRNA is now charged, because it is holding onto an amino acid thanks to aminoacyltRNA synthetases 4. uncharged: tRNA without an amino acid IV. Initiation of Translation a. Initiation: components assemble (mRNA, small and large subunits of ribosome, initiation factors, initiator tRNA and GTP) b. Three major steps: i. mRNA binds to small ribosomal subunit ii. initiator tRNA binds to mRNA iii. Large ribosome subunit joins initiator complex c. Initiation Factor 3 (IF3): binds to the small subunit to keep the large subunit away. i. allows for the small subunit to attach to the mRNA ii. helps the small subunit to find the ShineDalgarno sequence 1. ShinDalgarno sequence: in bacteria transcripts. Essential in transcription a. Looks similar in every bacteria, can have some base changes b. Interacts with rRNA of small subunit c. Can form hydrogen bonds d. Attaches small subunit in the correct position (three bases away from the start codon) i. Start codon: AUG ii. ShineDalgarno sequence is always in the same spot from the start codon iii. That way the start codon will be lined up in what will become the P site of the ribosome fMet d. fMettRNA : i. the first fMet (the one before the –tRNA) says that it is a charged tRNA that is holding onto fMet. The second fMet (the one that tRNA is raised to) says that this tRNA is supposed to be charged by fMet fMet ii. uncharged: tRNA iii. if messed up and charged incorrectly, it could look like this: lystRNAet 1. will read for fMet and put down lys iv. fMet is special nformyl methamine**** 1. prokaryotes have fMet for initiation, after that it will use regular Met 2. Eukaryotes have normal Met 3. If you see fMet, then you know it is a prokaryote V. Elongation a. Requires: i. 70s or 80s complex (the ribosomal complex, 70s=prokaryotes and 80s=eukaryotes) ii. Charged tRNAs iii. Elongation factors, GTP b. Initiator tRNA binds in the P site i. Only site for initiator tRNA to bind to c. During elongation, all tRNA will bind to the A site, then P site, then E site i. APE d. A, P, and E sites are all three bases wide e. Peptidyl transferase: cuts amino acids in the P site, pastes amino acid in a site, forms peptide bond formation when it is pasted i. rRNA component, catalytic RNA or ribosome; carries out this activity f. The process: i. New charged tRNA goes into A site ii. Cuts amino acid from P site and then pastes it to A site iii. Scoots down or translocate iv. Repeats v. Peptide bonds form between amino acids vi. Creates polypeptide (connection of all of the amino acids) g. Ribosomes read RNA 5’ to 3’ i. Ribosome starts at the ShineDalgarno sequence ii. Reads 5’ to 3’ every three bases iii. Insertion or deletion will not stop this system from working, but will affect the amino acid placed h. Termination: i. No tRNA matches the stop codons 1. Instead of tRNA, a release factor will fill the A site, will get cut and tried to be pasted, but will not be able to. This will stop the process 2. Release factor will float away and the subunits break apart, and releases the polypeptide chain. VI. Prokaryote vs. Eukaryote a. Genetic Code: This is conversed i. Prokaryotes: fMet is an initiator amino acid. To start, fMet will be first placed into the P site to help align the start codon ii. Eukaryotes: do not have fMet. A normal Met is used as an initiator to make sure the start codon is placed in the right spot. A specialty tRNA is used to get everything started. b. Transcription and translation occur simultaneously in prokaryotes i. Every time the ShineDalgarno sequence is exposed, another ribosome will attach c. mRNA life Span: i. Prokaryotes: short lived 1. Due to transcription and translation being able to occur in same place ii. Eukaryotes: live longer 1. Due to transcription and translation being unable to occur in the same place d. Ribosomal Subunits: differences create differences in initiation i. Prokaryotes: smaller in size overall, still two subunits ii. Eukaryotes: larger in size overall, still two subunits VII. onegene, oneenzyme/onegene, onepolypeptide a. Garrod: suggested that genes are encoded enzymes (first to suggest this) i. was studying what we know now to be genetic diseases ii. noticed that they were being passed down within families from one generation to the next iii. thought this was due to a protein not working correctly 1. made a connection between heredity and proteins, this was long before DNA was known as genetic material b. Beadle and Tatum: experimentally proved what Garrod was trying to say i. Used bread mold, easy to grow in lab setting and the vegetative form is haploid (can see recessive mutations easily) ii. They were the first to make the connection that if you mutate genetic material, then you will cause a gene to not be present, which leads to an enzyme not being created, which will lead to a phenotype missing iii. Theory: Onegene, oneenzyme. Each gene encodes a separate enzyme 1. later changed to onegene, onepolypeptide a. sometimes many polypeptides are used to interact to create a functional product iv. The experiment: 1. Exposed spores to UV radiation a. Will cause spores to mutate b. Done in complete media 2. Switched spores to minimal media a. Looked to see if the spores could grow or not b. Have to have the wild type allele to be able to live, if the spores can make it on their own and grow in minimal media, then they did not mutate c. If they die, then they had mutated d. All died 3. Then switched to supplemental media to see what the spores needed to live, to see what they are mutant for a. If they live in a certain media, then that is what they are missing, what they are deficient in b. The experiment showed that they grew in media with amino acids added. This means that they cannot grow an amino acid 4. They then put the spores in media with a certain amino acid to see what amino acid they cannot make a. Discovered it was tyrosine 5. Conclusion: change the gene, change the amino acid, change the polypeptide VIII. Proteins: are needed and need to be made correctly a. Function: tied to structure. Can be enzymes, structural, signal transducers, other, or a combination of the listed. b. Structures: i. Primary: the actual sequence made during translation, the polypeptide chain ii. Secondary: interactions between neighboring amino acids, usually hydrogen bonds. This is the first interaction after being formed iii. Tertiary: forms the 3D structure of the protein. Results from interactions between secondary structures. 1. Can be functional in this structure, or moves to another structure iv. Quaternary: multiple polypeptides interacting c. Amino acids group i. Proteins are made up of 20 common amino acids ii. Amino acids are similar in structure: core of carbon, amino and carboxyl groups, and R group (which gives the amino acid its own characteristics and identification) iii. Broken down into groups: 1. Nonpolar: hydrophobic 2. Polar: hydrophilic 3. Polar: positively charged (basic) 4. Polar: negatively charged (acidic) d. Directionality: i. The amino group and carboxyl group will give directionality 1. Amino end is the first amino acid 2. Whatever is closest to the stop codon is the carboxyl end e. Modifications: i. Molecular chaperons: proteins start to fold as soon as they leave the ribosome (they are more stable folded). A molecular chaperon can come and hold off folding until a certain point, thus allowing the protein to form the correct shape. ii. Signal sequence: other proteins will recognize and put the signal sequence where they need to go. Once they are there, they will cut off that sequence and allow the protein to fold into the correct shape f. Prions: i. Prusiner: discovered that Scrapie in sheep was caused by infectious proteins ii. Some proteins fold and get transcribed correctly but for some reason will change configuration 1. This happens with prions. A regular protein is made correctly, but some change will occur. This results in a prion (most are neurologically related, located in the brain) to switch 2. Examples: Scrapie in sheep. Kuru in humans 3. If it switches, it will have the exact same RNA, DNA, and amino acids, but will have a different folding than the normal one. 4. Kuru: if normal protein bumps into a misfolded one (the prion), the normal will change into the prion. This will continue to bound into others, causing more prions to form Chapter 14: Gene Mutation, DNA Repair, and Transposition I. Mutations: can impact life in several ways a. Mutation: change at DNA level in genetic information that will be passed onto the next generation i. can be passed into daughter cells through mitosis or individuals through sexual reproduction b. can be good, allows for diversity, though most mutations are bad. c. We can use mutations to understand…. i. how different genes function ii. Where genes are located iii. When something goes wrong in a gene, what affect does it have on phenotype II. Somatic Mutations vs. GermLine Mutations a. Tissue type change: i. Does it occur in somatic tissue or germline? 1. Somatic mutations: arise in somatic (nonsex) cells. They are passed to daughter cells through mitosis a. They divide mitotically b. Through mitosis, the daughter cells can have the mutation, but just on a region of the skin surface c. Will not be super dramatic, can be cancerous if it occurs in the wrong tissue 2. Germline mutations: arise in cells that produce gametes and are passed to future generations a. Occur in tissues that are passed onto the next generation of an organism b. Passed onto a brand new individual 3. Germline will result in the entire organism having mutations, not just in a region III. Classifications of Mutations: a. These classifications are not mutually exclusive, can be classified in multiple classifications b. Spontaneous mutations vs. induced mutations i. Spontaneous mutations: occur without a real cause, nothing is interfering with DNA to cause the mutation 1. Examples: replication errors that are not caught or fixed ii. Induced mutations: outside factors influence/causes the mutation 1. Naturally induced: is something we cannot escape from that causes the mutation. Such as UV light 2. Artificially induced: a chemical that results in a change at nucleotide level, usually something that can be avoided. iii. Base/Base pair substitution or Point Mutation: a single nucleotide is altered. Occurs at the sequence level 1. Transition: replaces a like for a like. a. a purine replaces a purine b. a pyrimidine replaces a pyrimidine 2. Transversion: replaces unalike bases a. a purine replaces a pyrimidine b. a pyrimidine replaces a purine iv. insertion/Deletion: the most common type 1. Inseriton: inserting a new base 2. Deletion: deleting a base 3. Any time you add or take out bases, you change the reading frame a. Frameshift mutations will occur v. Frameshift mutations: 1. Detrimental 2. If had to choose between point mutation and frameshift mutation, choose the point! 3. Point mutation: only going to change the exact location where the mutation is put or switched out a. It is a single point that is mutated 4. Deletion/insertion: everything after the point of deletion or insertion will be messed up a. The rest of the polypeptide will be changed vi. Outcomes from a mutation: 1. When talking about mutations, it is usually talking about forward mutations 2. Forward mutations: wild allele is changed to a mutant allele a. Missense mutation: single base changed that results in the incorporation of a different amino acid b. Nonsense mutation: single base change that results in a stop codon being created i. Stops the process of translation, loss of material c. Silent mutation: changes the base resulting in a different codon, but the same amino acid due to the wobble effect 3. Reverse mutations: goes from mutant allele to a wild allele a. A true reverse mutation will undo whatever forward mutation took place 4. Neutral change: changes amino acid, but not how the protein works 5. Loss of function mutation: prevents protein from doing the job a. Null: complete loss of function b. Haploinsufficiency: protein works, but not as well 6. Gain of function mutation: production of a new trait, protein takes on a new job 7. Suppressor mutation: hides the impact of another mutation. Returns to wild type phenotype but will not fix the mutation, hides the mutation with a second mutation a. Intergenic mutation: the second mutation occurs in a separate gene b. Intragenic mutation: the second mutation occurs in the same gene IV. Mutagens: a. Chemical mutagens: this means that we are not going to have damage at the DNA level caused by a specific chemical b. There are a lot of different types of mutagens i. Alkylation: adding methyl and ethyl groups to bases ii. Deamination: removing main groups of bases iii. Hydroxylation: adding in hydroxyl groups to bases iv. Understand that by modifying/changing these bases, even if just for a moment, especially during replication, can change what will be there permanently. The more chemicals exposed to the more changes created c. Intercalating agents: are mutagenic i. Intercalation: not going to modify bases, but insert themselves into the DNA and will stick between the bases ii. Toxic for the cell iii. As any machinery tries to work through the DNA, these will block the process 1. If present during replication and not fixed, it will cause insertions and deletions iv. Used in the lab to visualize DNA d. Radiation: can be mutagenic i. Can be considered artificial or natural ii. Wavelength will determine how much damage it will do, the more concentrated the wavelength, the more damage it can do to cells and DNA iii. We often focus on UV, it has less energy than other rays, but the bases will absorb this energy and can cause changes at the DNA level iv. Most common is pyrimidine dimers 1. Occurs mostly in T 2. Instead of hydrogen bonds, it will form covalent bonds with neighboring T. this will distort the backbone 3. Too many dimers will stop polymerase and cause death 4. Exposing equipment to UV light, we can kill cells V. Repair: a. Mismatch: i. Will occur right after replication takes place ii. When there is a base that is wrong, a mismatch, it will distort the DNA and this mismatch machinery recognizes the mistake and knows it needs to fix it iii. Replication is completed, so which strand needs to be fixed? 1. In bacteria: bacteria will methyl their DNA on both strands. Right after replication, the new strand has not been methylated yet. The machinery will search for methyl and will know that that strand is the old strand and will fix the new strand. a. will nick the new strand at the area of the methyl of the old strand, chew out and remove all of the DNA until it reaches the mistake, repair polymerase will come and fill in the gap of the old strand 2. Eukaryotes: do not mark their strands with methyl, makes it harder b. Direct DNA repair i. Does not replace nucleotides, it is going to correct modified bases ii. If we have a guanine with a methyl group, we remove the methyl group to create a normal guanine iii. Same base, just removal of modification c. Base Excision Repair: excises modified bases i. But out a base ii. Going to be used to correct a single damaged base iii. We will see a damaged base, go in and remove the base, leave behind the sugar and phosphate, use endonuclease to remove the backbone to leave a hole in the DNA iv. DNA polymerase will come in and read what base is on the template, fill it in, ligase will come and seal it to the backbone d. Nucleotide Excision Repair i. One of the most important ii. Recognizes a lot of different types of damage, focuses on large lesions iii. Recognizes damaged DNA and binds to region, removes multiple nucleotides iv. Unwinds in the region, use single stranded binding proteins to keep it single stranded/unwound v. Will cause a cut on either side of the region 1. Gets ride of a whole chunk 2. Removes it, leaving behind single stranded template, polymerase comes in to fill it and ligase will seal it e. Homologous Recombination i. Occurs after replication has taken place. At the end or near the end of S phase or near G2 phase ii. Have the ability of using sister chromatid as a template iii. Machinery recognizes the mistake of the chromatid. Will chew back some and leave overhands of the one with the break iv. Will look for complementary part in the sister chromatid and will bind the them together and use the sister as a template v. This allows for repair without losing any genetic information vi. Homologous recombination repair allows for fairly error free fixing of DNA f. Nonhomologous end joining: i. Error prone ii. No sister chromatid is present. iii. Will grab the ends of the damaged parts and stick the ends back together. iv. Results in the loss of genetic information VI. Ames Test a. What it does: determines if a chemical or compound is mutagenic i. Mutagenic: does not guarantee it is carcinogenic, but there is a high possibility b. Uses auxotroph: mutant bacteria c. Uses His bacteria: bacteria that cannot make His, if in a media that does not have His, it will die. His is used as a marker, follow and trace to see if anything changes at DNA level d. How we carry out the test: i. Use liver enzymes: some chemicals are okay until we try to break them down in the liver which can activate chemicals or compounds ii. Mix bacteria with liver enzymes: place it in His media 1. Control: see how many revert back normally (the rate spontaneously changes back) iii. Take another sample and repeat above, and mix with chemical being tested 1. Treatment plate: do we have more colonies (reversions) then the control? a. If the same: the chemical does not cause damage b. If more: the chemical does cause mutations i. Is a mutagen, can be carcinogenic e. Conclusion: for the cell to grow in His media then it has revert back and become his+. If it grows in His, then there was a change (mutation), meaning the chemical is mutagenic VII. Transposable Elements: a. Repetitive and mobile. Can also cause mutations b. 50% of genome c. Common features: i. Terminal inverted repeats inside the element 1. Allows for them to be cut out of DNA and placed back in DNA ii. Flanking: won’t move, they are outside of the element d. Code for own gene product, allows for gene to be cut out and inserted elsewhere e. Transpositions: i. Creates staggered cuts and creates overhangs ii. Elements will place itself in and then DNA repair mechanisms will fill in the gaps, and seal the transposable element into the DNA iii. When filling in gaps, it will create repeated portions: flanking repeats f. Barbara McClintock: discovered transposable elements i. Worked with maze, looked at pigmentation: purple pigment, no pigment, and variated pigment ii. Transposase allows for transposable elements to be cut and placed elsewhere iii. Ac Element: activators, they are autonomous, can take care of themselves 1. Make transposase gene: enzyme that is necessary for enzyme to be able to move and cut themselves out iv. Ds Element: nonautonomous, smaller than Ac. They are smaller because they had deletions in transposase gene, they cannot make active transposase and cannot jump on their own. 1. if transposase is made by Ac element, they can find the borders and help the Ds element jump 2. can only jump if a transposase is being made by Ac element v. cc is no pigment and Cc is purple pigment 1. when Ds element jumps into C of a Cc gene, it creates no pigment, when it jumps out, it will create the purple pigment again, causing a variated pigment VIII. Transposable Elements are Mutagenic a. Any time something can insert itself it can disrupt the flow of information i. If inserted at DNA level and then is translated it will create the wrong protein b. Hemophilia: there has been three separate families where a line has inserted itself into the gene that causes hemophilia c. BRAC2: breast cancer i. Alu d. Can be mutagenic caused by how many there are that exist Exam 4 Study Guide cont. Chapter 15: 1. Definitions: a. Benign: tumor that is localized, stays in one place b. Malignant: tumor that moves to other tissues, considered cancerous at this point c. Metastasis: tumor that moves to other sites, when breast cancer moves to different site, it is still considered breast cancer d. Apoptosis: cell death that is programmed, cell will sacrifice itself because it knows something is not right e. Oncogene: mutated copy of gene f. Tumor suppressor genes: push cells into apoptosis, tries to reduce cancerous cells g. Pseudodominance: appearance of a trait in a cell that is heterozygous for a normally recessive trait h. Telomerase: activated in many tumor cells, allows unlimited cell division i. Vascularization: tumor cells need nutrients j. miRNA: controls gene expression k. Epigenetic: changes in methylation/acetylation of DNA/histones 2. Multihit hypothesis: a. Alfred Knudson: several mutations have to occur before a cell can become cancer i. Broke his population into two groups when dealing with eye cancer: 1. Older patients: get cancer in one eye, no relative history to the cancer 2. Younger patients: often get cancer in both eyes, relative history with the cancer 3. Hypothesis: have to accumulate multiple mutations to allow for progression to cancer. ii. Clonal evolution: multiple rounds allow tumor cells to become aggressive 1. Repair mechanisms: are in place to prevent the cell from further dividing 2. Apoptosis: cell is programmed to die if something is not right
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