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This 7 page Class Notes was uploaded by Shira Clements on Monday March 28, 2016. The Class Notes belongs to BSCI105 at University of Maryland taught by Norma Allewell in Fall 2015. Since its upload, it has received 21 views. For similar materials see Principles of Biology I in Biology at University of Maryland.
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Date Created: 03/28/16
Shira Clements BSCI105 Chapter 16- Molecular Basis of Inheritance The Search for Genetic Material (DNA) - Early in 20 century, identifying molecules of inheritance was a major challenge - Morgan showed that gene exists as part of chromosomes (made of DNA and proteins), so they were tested on for genetic material o Before scientists were testing proteins, but it came out as different results, so they knew they were wrong, so when Morgan and Mendel figured it out, it was revolutionary o Worked on bacteria to find out this information Evidence DNA Can Transform Bacteria - 1928- attempt to make vaccine against pneumonia, Griffith was studying the bacteria that causes pneumonia in mammals o Had two strains of bacteria- one pathogenic (causes disease) and other non-pathogenic (harmless) Surprising- when he killed the pathogenic bacteria of nonpathogenic strain, some of the living cells became pathogenic, so that means that pathogenicity was inherited by decedents of the bacteria. Clearly some chemical of the dead pathogenic cell caused the heritable change Transformation- change in genotype and phenotype because of assimilation of external DNA by a cell. - Avery was now inspired o Focused on DNA, RNA (other nucleic acid in cell), and proteins o Broke open the heat killed pathogenic bacteria and took cellular contents o Took DNA, RNA, and protein and inactivated one of them and tested its ability to transform live nonpathogenic bacteria Only when DNA was active, the transformation occurred Hence, in 1944, it was announced that DNA is the transforming agent, but many scientists still doubted it Evidence that Viral DNA Can Program Cells - More evidence that DNA was the genetic material came from studies that infect bacteria= bacteriophages (aka phage) - Virus- is just a little more than DNA with a protective coat around it, mostly a protein o to produce viruses, the virus must infect a cell and take over the metabolic machinery - 1952- Hershey and Chase showed that DNA is genetic material of phage (T2) which infects e coli, so they experimented on e coli o At that time, they knew that T2 was made up mostly DNA and protein and that T2 can infect e coli and make it release many more T2 when it breaks, but needed to find out if it was protein or DNA Used radioactive isotope of sulfur to tag to protein in one and DNA in another, and they found out that the phage DNA entered the host cell but phage protein did not. With the DNA, it let out many others with the virus. o Concluded that DNA is the molecule to carrying genetic information that makes cell produce new viral DNA and protein. – clear evidence that it is DNA and not protein Additional Evidence that DNA is the Genetic Material - Chargaf- already known that DNA is polymer of nucleotides, each having a nitrogenous base (A, T, G, or C), pentose sugar of deoxyribose, and a phosphate group Rules that were explain o Base composition varies from one species to anotherwhen double helix came o Number of A=T and number of G=C along Building Structural Model of DNA - Now- how could structure fit role of inheritance - 1950s- arrangement of covalent bonds in nucleic ace polymer was known - 1953- Watson and Crick- used X-ray crystallography and studied protein structure. Watson saw an Xray defraction picture of DNA produced by Franklin, and a picture produced by xrays that were deflected as they passed through aligned fibers of DNA, which were both helical because of defractions, which just confirmed his beliefs that DNA was helical- made up of two strands= double helix. o Started to make models of double helix which would conform to xray measurements and the known chemistry of DNA (including Chargaff and with Franklin’s-female- information saying that sugar phosphate backbones were on outside of DNA) Hydrophobic parts were in interior and nitrogenous Like twisted bases on interior ladder Negatives charges weren’t near each other Sugar phosphates were antiparallel- subunits run in opp direction A-T, and G-C (need a purine and pyrimidine to be right size) DNA Replication and Repair Basic Principle- Base Pairing to a Template Strand - Watson and Crick hypothesis- two strands that are complementary to each other o Before it duplicates, hydrogen bonds are broken, and two chains unwind and separate, and then each acts as template for new chains, so it would be 2 exact same strands Nucleotides just match up to complementary strand Each daughter molecule will have one old parental strand and one new strand- semiconservative model compared to conservative model where both strands just come back together after process and compared to dispersive model where all four strands have some new and some old Meselson and Stahl confirmed the semiconservative model DNA Replication - E coli has one chromosome with 4.6 million nucleotide pairs, and can replicate this DNA in less than an hour, but we have 46 DNA molecules in each cell in nucleus with long double helix in chromosome, which in all has 6 billion nucleotide pairs. - Proteins and enzymes help it along - Step 1- begins at origins of replication (short stretches of DNA with a specific sequence of nucleotides), and then proteins attach to the DNA there and separate the strands making a replication bubble. Replication proceeds in both directions until entire molecule is copied. o Bacterial chromosomes only have one origin, while eukaryotic have many- multiple bubbles form with each other and fuse together, speeding up process Both- replication proceeds in both directions from each origin o At end of replication bubble is a replication fork- Y shaped region where parental strands of DNA are unwound (the untwisting, twists the twisted part even more- pulls it tighter) Helicases- enzymes that untwist the double helix at replication forks, making them available to be a template strand Single strand binding proteins- bind unpaired DNA strands, keeping them from repairing with each other Topoisomerase- helps relieve tightening the twisted strands by breaking, swiveling, and rejoining DNA strands. o Now a template, but enzymes that synthesize DNA cannot start synthesis of polynucleotide, rather they can only add nucleotides to the end of existing chain that is base-paired with template. Initial nucleotide chain that is produced during DNA synthesis is rally RNA= primer and synthesized by primase, starts complementary RNA strand from single RNA nucleotide, adding RNA nucleotides one at a time, using the DNA template, which is then base paired to template and will start from 3’ end of RNA primer. - Step 2- Enzyme DNA polymerase- catalyze synthesis of new DNA by adding nucleotides to preexisting chain- bacteria have 2 while eukaryotes have 11 o Require primer and DNA template strand, and add DNA nucleotides to RNA primer and then continue to DNA strand o Each nucleotide added becomes a nucleoside triphosphate (sugar and base with three phosphate), just like ATP but with deoxyribose instead of ribose, so dATP that supplies adenine nucleotide to DNA Chemically reactive because triphosphate tail has unstable cluster of neg charge As each monomer joins, two phosphate groups are lost as pyrophosphate, which is the hydrolyzed with two molecules of inorganic phosphate and is a coupled exergonic reaction that helps drive the polymerization reaction o 2 new strands are antiparallel to the template too Primases can only add to the free 3’ end of primer or growing DNA strand, not 5’ end. Therefore, new DNA can only elongate in 5’ to 3’ direction Leading Strand- DNA strand made by adding nucleotides (the complementary strand) Lagging Strand- the other DNA strand is the mandatory 5’ to 3’ direction, so DNA polymerase must work along the template strand in direction away from the replication fork because can only add in 5’ to 3’, but has to be antiparallel It adds nucleotides discontinuously, in segments= Okazaki fragments 1. Primase joins RNA nucleotides into primer DNA polymerase III adds DNA nucleotides to primer of fragment 1 After it touches the next primer, polymerase detaches Fragment 2 is primed, and DNA polymerase adds DNA nucleotides, detaching when it reaches the fragment 1 primer DNA polymerase I replaces RNA with DNA, adding 3’ end of fragment 2 DNA ligase forms a bond between the newest DNA and the DNA fragment 1- joins final nucleotide of replacement DNA to first DNA nucleotide of adjacent Okazaki fragment The lagging strand in this region is now complete o DNA polymerases are represented as trains moving down a track, which is wrong because: Many proteins interactions facilitate form a large DNA complex Primase acts as a brake at the fork when other proteins interact with it and slows down the process and coordinates the placement of primers and rate of replication on both strands DNA replication complex may not move, rather the DNA moves through the complex- lagging strand loops back around through the complex DNA Proofreading and Repairing - DNA polymerases proofread each nucleotide with template- if incorrect, then it will remove the nucleotide and resume to synthesis - Mismatch Pair- other enzymes remove and replace incorrectly matched enzymes o This pair can cause cancer - Incorrectly or altered pairs- can damage DNA if not repaired, bad pairs happen often, but usually fixed before permanent; if they aren’t, it is called a mutation o Many enzymes are present to try to fix them o Most use mechanism that takes advantage of the base-paired structure of the DNA Nuclease- the DNA cutting enzyme that takes out incorrect pair, then the resulting gap is filled with other nucleotides via DNA polymerase or DNA ligase, using undamaged strand as template- this whole thing is called nucleotide excision repair Replicating the Ends of DNA Molecules - Because DNA polymerase can only add to the 3’ or a preexisting template, there is no way that it can complete the full replication of a daughter DNA strand. Even if it is started with RNA primer, the primer is removed, and then it cannot be added before because of the 5’ end; therefore, repeated rounds of replication produce short DNA molecules with uneven/staggered ends. Therefore, eukaryotic chromosomal DNA molecules have special nucleotide sequences called telomeres at end. o Telomeres don’t have genes, but repeat certain nucleotide sequence many times, which then protects the organisms gene o Proteins associated with telomeres prevent staggered ends of daughter cells from activating, which prevents damage o They become shorter during every round of replication o Telomerase catalyzes the lengthening of telomeres so genes are not lost and chromosomes are not shortened o Shortening of telomeres can protect organisms from cancer by limiting divisions that somatic cells can undergo Cells from large tumors have short telomeres, since it underwent cell divisions, and further shortenings would lead to self destruction of tumor cells, but telomerase would allow these cancer cells to persist because it wouldn’t make the telomerase shorter, so telomerase is imp in cancer cells - Prokaryotes have circular chromosome so shortening does not occur Chromosome - Chromosome- consists of DNA molecule packed together with proteins o They are coiled- densely packed in there, so it is really a lot bigger than cell if it was stretched out o In bacteria it is in the nucleoid- which is a not bounded membrane o In eukaryotic cells- single DNA double helci averages about 1.5 x10 nucleotide pairs and would be 4 cm if stretched out In Eukaryotic cells, DNA is combined with protein, which together make make up a complex called chromatin, which fits into the nucleus. Chromatin Packing in Eukaryotic Chromosomes - DNA and the phosphate groups have neg charge, which is on the outside of each strand - Histones- small proteins that are responsible for first level of DNA packing in chromatin o Positively charged so bond to DNA o DNA wraps around, so it structures and orders DNA into nucleosomes - Nucleosome- basic unit of DNA packing o each individual wrapping, the individual bead on a string o DNA wound twice around a protein core composed of two molecules of each of the four main histones, and the histone tail (N terminus) is facing outward - Interactions between histone tails and linker DNA and nucleosomes on either side o 5 histone is involved o the interactions cause the fiber to coil, forming a chromatin fiber with 30nm in thickness - The fiber then forms a loop called looped domain, attached to a chromosome scaffold made of protein (together=300nm fiber) - In mitotic chromosome- looped domain coil and fold, which compacts all chromatin, which then makes the classic chromosome picture of 1400nm (each chromatid 700nm)
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