Chapter 10 The Molecular Basis of Inheritance Lecture Outline Overview: Life s Operating Instructions

Transcription

1 Chapter 10 The Molecular Basis of Inheritance Lecture Outline Overview: Life s Operating Instructions In April 1953, James Watson and Francis Crick shook the scientific world with an elegant double helical model for the structure of deoxyribonucleic acid, or NA. Your genetic endowment is your NA, contained in the 46 chromosomes you inherited from your parents and the mitochondria you inherited from your mother. Nucleic acids are unique in their ability to direct their own replication. The resemblance of offspring to their parents depends on the precise replication of NA and its transmission from one generation to the next. It is this NA program that directs the development of your biochemical, anatomical, physiological, and (to some extent) behavioral traits. Concept 10.1 NA is the genetic material. The search for genetic material led to NA. After T. H. Morgan's group showed that genes are located on chromosomes, the two constituents of chromosomesproteins and NA, became the candidates for the genetic material. The Work of Frederick Griffith Until the 1940s, the great heterogeneity and specificity of function of proteins seemed to indicate that proteins were the genetic material. However, this was not consistent with experiments with microorganisms, such as bacteria and viruses. The discovery of the genetic role of NA began with research by Frederick Griffith in Griffith studied Streptococcus pneumoniae, a bacterium that causes pneumonia in mammals. One strain was harmless. The other strain was pathogenic (disease causing). Griffith mixed a heat killed, pathogenic strain with a live, harmless strain of bacteria and injected this into a mouse. The mouse died, and Griffith recovered the pathogenic strain from the mouse s blood. Griffith called this phenomenon transformation, a phenomenon now defined as a change in genotype and phenotype due to the assimilation of external NA by a cell 1

2 2

3 Work of Oswald Avery For the next 14 years, American bacteriologist Oswald Avery tried to identify the transforming substance. Avery focused on the three main candidates: NA, RNA, and protein. Avery broke open the heat killed, pathogenic bacteria and extracted the cellular contents. He used specific treatments that inactivated each of the three types of molecules. He then tested the ability of each sample to transform harmless bacteria. Only NA was able to bring about transformation. Finally, in 1944, Oswald Avery, Maclyn McCarty, and Colin MacLeod announced that the transforming substance was NA. Still, many biologists were skeptical because proteins were considered better candidates for the genetic material. There was also a belief that the genes of bacteria could not be similar in composition and function to those of more complex organisms. Further evidence that NA was the genetic material came from studies that tracked the infection of bacteria by viruses. Viruses consist of NA (or sometimes RNA) enclosed in a protective coat of protein. o To replicate, a virus infects a host cell and takes over the cell s metabolic machinery. o Viruses that specifically attack bacteria are called bacteriophages or just phages. 3

4 Confirmation of NA as Genetic Molecule by Hershey and Chase In 1952, Alfred Hershey and Martha Chase showed that NA was the genetic material of the phage T2. The T2 phage, consisting almost entirely of NA and protein, attacks Escherichia coli (E. coli), a common intestinal bacteria of mammals. This phage can quickly turn an E. coli cell into a T2 producing factory that releases phages when the cell ruptures. To determine the source of genetic material in the phage, Hershey and Chase designed an experiment in which they could label protein and NA and then track which entered the E. coli cell during infection. o They grew one batch of T2 phage in the presence of radioactive sulfur, marking the proteins but not NA. o They grew another batch in the presence of radioactive phosphorus, marking the NA but not proteins. o They allowed each batch to infect separate E. coli cultures. o Shortly after the onset of infection, Hershey and Chase spun the cultured infected cells in a blender, shaking loose any parts of the phage that remained outside the bacteria. o The mixtures were spun in a centrifuge, which separated the heavier bacterial cells in the pellet from the lighter free phages and parts of phages in the liquid supernatant. o They then tested the pellet and supernatant of the separate treatments for the presence of radioactivity. Hershey and Chase found that when the bacteria had been infected with T2 phages that contained radiolabeled proteins, most of the radioactivity was in the supernatant that contained phage particles, not in the pellet with the bacteria. When they examined the bacterial cultures with T2 phage that had radiolabeled NA, most of the radioactivity was in the pellet with the bacteria. Hershey and Chase concluded that the injected NA of the phage provides the genetic information that makes the infected cells produce new viral NA and proteins to assemble into new viruses. The fact that a cell doubles its amount of NA prior to mitosis and then distributes the NA equally to each daughter cell provided some circumstantial evidence that NA was the genetic material in eukaryotes. Similar circumstantial evidence came from the observation that diploid sets of chromosomes have twice as much NA as the haploid sets in gametes of the same organism 4

5 Bacteria hage attacking E. Coli bacteria Hershey Chase Experiment 5

6 Chargaff's Rule (Edwin Chargaff) By 1947, Erwin Chargaff had developed a series of rules based on a survey of NA composition in organisms. o o He already knew that NA was a polymer of nucleotides consisting of a nitrogenous base, deoxyribose, and a phosphate group. The bases could be adenine (A), thymine (T), guanine (G), or cytosine (C). Chargaff noted that the NA composition varies from species to species. In any one species, the four bases are found in characteristic, but not necessarily equal, ratios. He also found a peculiar regularity in the ratios of nucleotide bases, known as Chargaff s rules. In all organisms, the number of adenines is approximately equal to the number of thymines (%T = %A). The number of guanines is approximately equal to the number of cytosines (%G = %C). Human NA is 30.3% adenine, 30.3% thymine, 19.5% guanine, and 19.9% cytosine. The basis for these rules remained unexplained until the discovery of the double helix. 6

7 10.3 Structure of NA determines its function/ NA structure From all of this evidence a two dimensional structure was created: 7

8 Creating the three dimensional model of NA: Rosalind Franklin: This hotograph served as this starting point for Watson and Crick: Chargaff's rule and Franklin's photo allowed Watson and crick to determine the correct 3 dimensional structure Base airing: Complementary bases form hydrogen bonds to keep the two strands together urines yrimidines 8

9 10.4 NA Structure and Replication A. NA Structure " ouble Helix" 2 Main Areas: 1. Backbone hosphates Groups eoxyribose Sugar 2. Rungs= Nitrogenous Bases Adenine (A) A Guanine (G) G Cytosine (C) C Thymine (T) T These things form the monomers of NA= A Nucleotide Backbone Rung A Nucleotide Adenine NA is ouble stranded the two strands are held together by a weak hydrogen bond Base airing: 5' 3 ' G C 3 ' 5' A always base pairs with T G always base pairs with C T A Two strands are Antiparallel A T C G T A 5' 3 ' G C 3 ' 5' 9

10 NA Replication A. Is done one Chromosome at a time Eukaryotes have linear chromosomes rokaryotes (Bacteria) have circular chromosomes called plasmids B. A Chromosome contains: Nucleosomes which are 1. Long supercoiled segments of NA allows a lot of NA to fit into a small space. 2. roteins called Histones responsible for keeping NA coiled lasmid 10

11 11

12 B. NA replication happens according to the semiconservative model 12

13 C. Simplified Overview of Replication Replicated NA contains an original strand (Template Strand) and a newly synthesized complementary strand (aughter Strand) NA Replication does not begin at one end and end at the other Happens at many locations at one time known as Replication Bubbles 13

14 . Enzymes Involved in NA Replication 14

15 E. Synthesis and Elongation of new NA strand (Helicase and NA olymerases) 1. NA is unwound by the Enzyme Helicase in both directions creating the Replication Bubble 2. Synthesis of the aughter strand always takes place in the 5' to 3' irection 3. Replication begins in the center of the bubble Origin of Replication in both directions a. this creates the leading strand and the lagging strand 1. Leading strand moves forward towards Replication fork 2. Lagging Strand moves backwards away from Replication Fork 15

16 Synthesis of the Leading and Lagging Strand Must progress in the 5' to 3' direction 16

17 17

18 F. NA synthesis must be rimed before proceeding 1. Because NA olymerase III (oly III) can only join nucleotides to the 3' end of the growing chain, it cannot initiate copying 2. This is done by using a rimer a short sequence with an available 3' end (usually RNA) a. the primer is built by rimase b. rimer is later replaced by NA (oly I) c. Leading strand only needs one primer d. Lagging Strand Each Okazaki fragment needs its own primer 18

19 Overview of NA Replication 19

20 20

21 G. NA Must be proofread and mistakes repaired 1. Synthesis results in typos ( 1 in 100,000 B) 2. These will result in mutation if left unfixed 3. Mistaken segments are cut out by Nuclease in Nucleotide Excision Repair 21

22 4. art of this roofreading process allows NA to replace its rimers 5. However this results in shorter and shorter NA molecules To avoid loss of genetic information, each NA molecule is capped by a genetically meaningless segment called a Telomere 22

23 Telomeres indicated in orange segments at end of chromosome 23

24 24