jimtrue.com : school : BSC2010 : CH 16: Molecular Basis of Inheritance

Posted by Jim True on April 15, 2004 6:31 AM. Last Updated October 22, 2006 9:23 PM

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CH 16: Molecular Basis of Inheritance

Nucleic Acids

• Nucleic acids were first discovered in 1869.
• Because these molecules were found mainly in the nucleus of eukaryotic cells, and because the pH of the molecules in solution was slightly acidic, the molecule were called nucleic acid.
• In the 1920's, DNA and RNA were found to be different molecules.
• RNA - Ribonucleic Acid, and
• DNA - Deoxyibonucleic acid
• DNA and RNa were found to possess three different types of compounds:
  • Phosphate Group
  • 5 Carbon Sugar (deoxyribose in DNA, ribose in RNA).
  • Nitrogen containing ring compound (nitrogenous base).
• five different nitrogenous bases were identified:
  • Purine -- Two bases possessed double rings , Adenine (A) and Guanine (G)
  • Pyrimidine -- The other three bases possessed single rings, Thymine (T), Cytosine (C), and Uracil (U).
  • DNA has AGCT, RNA has AGCU.
• It was found that these three compounds formed a single uit called a nucleotide.
• Both DNa and RNA were formed by long chains of repeating nucleotides.
• Carbons in an organic molecule are numbered by organic chemists.
• A nucleotide is formed in such a way that the two functional groups are attached to different carbons:
  • Phosphate, attached to the sugar at carbon number 5 (called the 5' end of the nucleotide).
  • Hydroxyl, part of the sugar molecule at carbon number 3 (the 3' end).
  • Nucleotides are linked by dehydration sysnthesis betheween the hydroxyl and phosphate to form phosphodiester bonds.
  • Both DNa and RNa are linear molecules in eukaryotes, and so will always have an exposed phosphate group at one end (5' end) and an exposed hydroxyl at the other (3') end. (mnemonic: "five" - "phosphate").
  • this gives both molecules DIRECTION, ie. we can talk about a 3' end and 5' end and can examine processes that occur in specific directions.
  • When listing the sequence of nucleotides, we can indicate the direction of alignment by writing the 3' and 5' numbers at each end of the strand.
  • 5' N-N-N-N-3'
  • Because the phosphate group (P) is at teh 5 end and thydroxyl is on teh sugar (S) at the 3' end, we could write it as
  • 5' PSPSPSPS3'
  • the ONLY part of a nucleotide sequence that varies is the N base, because there are 4 different molecules.
  • The N base attached to the sugar at the number 1 carbon.
  • Because it is the bases that vary a linear strand of DNA or RNA only lists the base letters:
  • DNA 5' GCTATTC 3'
  • RNA 5' CCUAUUC 3'
  • Note that RNA has the N base Uracil instead of Thymine.

DNA as Genetic Material

• It was also during the 1920's that genes were first linked to chromosomes.
• Once this was shown, the two major components of the chromosomes, DNA and proteins, became the likely carriers of genes.
• Iniitally, proteins were thought to be the source of genes because they were so varied in structure and function.
• Gradually, attention shifted to DNA by studying extremly simple organisms such as bacteria, as well as viruses. Viral particles, is nothing more than a chain of nucleic acid (DNA or RNA) surrounded by a protein coat.
• Evidence from Bacteria:
  • Working with bacteria in 1928, Frederick Griffiths showed that bacteria can undergo transformation.
  • He worked with a bacterial species that had both disease causing (pathogenic "Agents of pathology - death or disease") and harmless strains.
  • Griffiths found that when heat-killed pathogenic bacteria were mixed into cultures of harmless ones, some of the live bacteria became pathogenic.
  • Experimental tests with both bacterial proteins and other compounds extracted frm dead bacteria and placed in living cell cultures showed that only DNa caused transformation.
  • Thus, transformation is the assimilation of external DNA released by dying bacteria into living cells.
• EVIDENCE FROM VIRUSES
  • Many viruses are particles containing a protein "coat" enclosing either DNA or RNA. (never both, or we've never found one that has both). Protein coat is shed by the virus when it is absorbed by the cell.
  • Viral "functions" are solely related to reproduction.
  • When viruses are attached to a living cell they take over cellular "machinery" and force the replication of new viral parts.
  • These are then assembled and destroy the host cell when the new viruses are released.
  • Viruses that infect bacteria are referred to as bacteriophages ("bacteria eaters").
  • In 1952, Alfred Hershey and marth Chase tested whether it was the protein or the enclosed DNA of bacteriophages that actually entered the cells.
  • They first "labeled" the viral proteins with radioactive sulfur and then allowed them to infect host cells.
  • Virtually no radioactive sulfur was found inside infected cells.
  • Radioactive phosphorus was used to label DNA and was found both inside bacteria and in newly created viral particles.
• Additional Evidence
  • DNA is doubled during S phase of Interphase and it is the DNa that is distributed equally to daughter cells during mitosis/cytokinesis or halved to gametes during meiosis.
  • It was originally thought that DNa was made up of a continuously repeating series of the 4 different bases: GCATCGATCGATGCAT ...
  • Thus, it was thought that all four bases occurred in equal quantitiies within a DNA molecule, ie. A = C = G = T.
  • In 1950, Erwin Chargaff discovered that this was not so. He found that within any DNA molecule, the quantity of A = T and G = C, but they did not equal each other.
  • This is now known as Chargaff's Rule, quantity of adenine is always equal to the quantity of thymine, cytosine is always equal to guanine.

DNA Structure

• In the 1950's research into the actual structure of the DNA molecule was intense.
• Working on DNA structure using a process called X-Ray diffraction, Rosalind Franklin and Maurice Wilkens added two more important facts on DNA structure.
• They showed that the DNA molecule forms in a spiral shape, and also provided precise dimensions to the diameter and distance between spirals.
• James Watson and Francis Crick are officially credited with providing the first accurate model of DNA, which they called a double helix model, in 1953.
• Double Helix Model:
  • Molecule is a spiral (helix) formed by TWO separate strands of nucleotides lying alongside one another.
  • The connections between the strands is formed by nitrogen base pairs.
  • To fit Franklin & Wilken's dimensions, a purine must bond to a pyrimidine.
  • Based on Chargaff's findings, it must be that A bonds to T, G bonds to C.
  • The bonds formed between the base pairs are hydrogen bonds!

DNA Replication

• There are three key characteristics that faciliatate the replication or repair of a DNA molecule:
  1. Complementary Base Pairing -- Either single strand (side) of DNA has any conceivable sequence of the four N bases.
     • The sequence of bases on the opposiste strand is exactly determined by the nature of these bases since
     • A always is matched to T, and G is always matched to C (Chargaff's Rule).
     • Thus, base pairs are complementary. Complement is to match, Compliment is to praise.
     • So if the first strand has AAATCGCGGC, then the second strand is TTTAGCGCCG
  2. Anti-Parallel Arrangement of Strands
     • In order for bases to line up so that they can bond across the strands, the strands are arranged with the exposed 3' and 5' ends ALIGNED in opposite directions.
     • 3' ATATATATAGC 5'
     • 5' TATATATATCG 3'
  3. Semi-Conservative Replication (or Nature of Synthesis)
     • Because of characteristics 1 and 2, in order to replicate any DNA molecule:
       • The helix only needs to be "unzipped" by breaking the H bonds at the base pairs.
       • Brand new complementary anti-parallel strands can be copied from each of the two original strands, ie, each original strand is a template for the formation of complementary anti-parallel duplicate strand.
     • Provides great accuracy in replication.
• Along both of the DNA strands are embedded series of nucleotides called initiation sites (origins of replication).
• At these locations, the process of DNa replication will begin, as the strands move apart, forming "replication bubbles".
• While DNA itself is a linear molecule in eukaryotes, DNA replication does not start at one end of the molecule and move linearly.
• Instead, it occurs at multiple locations simultaneously and works in both directions along the strands. (Figure 16.10, p. 295)
• The opening of the bubbles is accomplished by the attachment of a series of single strand binding proteins, keeping the single strands of the bubble separated.
• At each end of the bubble is a replication fork
• An enzyme called DNA helicase attaches to each fork and causes the bubble to continue to open in each direction.
• On each of the strands, complementary RNA nucleotides (aka Ribonucleotides) are brought to the strand at the initiation sites by an enzyme called primase.
• These join to form a short strand (~10 nucleotides long) called an RNA primer.
• The primer indicates the starting point for DNA strand replication.
• At this point, another enzyme, DNA polymerase, moves to the strands. DNA polymerase from sugar to phosphate end. 3' towards the 5' end.
• It was originally thought that the DNA polymerase moved along the DNA.
• It is now known that the DNA strand passes through the polymerase, bringing complementary DNa nucleotides (Deoxyribonucleotide) to the strand.
• However, DNa polymerase can ONLY pass the DNA strand through 3' --> 5'.
• Thus, the new strand produced by the polymerase will always be aligned in a 5' --> 3' direction.
• Leading Strands -- The portions of the strands that are aligned 3' --> 5' AWAY from the initiation site and TOWARDS the replication forks.
• Along leading strands, continuous synthesis of the new strands occurs, because the polymerase can continue to pull through this strand continuously as the replication fork unwinds the strands and smoothly add new DNA nucleotides.
• Lagging strands -- The portions of the strands that are aligned 3' --> 5' TOWARDS the initation site and AWAY from the replication forks.
• On these strands, after an RNA primer is set DNA polymerase begins to bring in DNA nucleotides and link them 5'--3', which moves the new strand pieces AWAY from the replication fork.
• As a result, the replication for moves AWAY from the new strands being formed on the lagging strands, creating a gap.
• In order to fill the gap, new RNA primers will have to produced by primases, and the DNa polymerases will need to "jump" back to the primers and begin again bringing in DNA nucleotides.
• This creates a series of DNA pieces.
• These short DNA pieces (100-1000 nucleotides) are Okazaki fragments.
• Thus, DNA replication on the lagging strand is termed discontinuous synthesis.
• The RNA primers between Okazaki fragments as well as between continuous segments will be stripped out and replaced by another DNA polymerase.
• Then, another enyzme, DNA ligase, links the sections by joining the unbonded sugars and phosphates (by dehydration synthesis).
• Replication bubbles form all along the DNA molecule and molecule and continue to develop in both directions until all the bubbles meet and join.
• At the end of the linear DNA in eukaryotes, there is a potential problem due to the inability of DNA polymerase to add nucleotides to the 5' end of a strand.
• How is it possible to do so?
• In prokaryotes, this is not a problem because the DNA forms a loop, but in eukaryotes, with linear DNA, there would be a progressive shortening of the DNA (and potential gene damage) because of this inability.
• Eukaryotic DNA have telomeres, "caps" on the ends of the DNA formed by a repeating sequence of nucleotides (100 -- 1,000 sequences). (Telo means "end", mere means 'part').
• Telomeres do NOT contain genes!
• Telomeres buffer the ends of the genetic DNA by being "eroded" ie shortening during successive DNA replications.
• Once erosion gets into genetic DNA, the cell will either die or stop dividing.
• Telomere length can be restored by the enzyme telomerase.
• Telomerase is present in multicellular organisms only in the earliest part of life when cells are dividing at a furious rate.
• They are not present in older cells which slow and stop dividing at a critical telomere length, thus telomeres may play a significant role in aging.
• However, telomerase has been found in cancer cells, which may suggest why cancer cells keep dividing and are essentially "immortal".

DNA Repair

• The intial copying of the old DNA strand by DNA polymerase results in about 1 in 10,000 copying errors.
• DNA polymerase then "proofreads" the strand and makes corrections, reducing the error count to about 1 in 1 BILLION!
• DNA is also constantly exposed to potential damage from numerous causes, e.g. radiation and chemical exposure.
• Damage often involves only a section of one of the two strands.
• Enzymes called nucleases cut out the damaged section and DNA polymerase attaches and repairs the affected section, using the undamaged strand as a template.
• This continuous process of proofreading and repair keep the rate of mutation (any alteration of a gene sequence) low. (Figure 16.17, p.299)
• Such repairs cannot be performed if there is massive chromosomal damage such as exposure to large doses of ionizing radiation. (or chemical exposure to teratogens).
• Such radiation can tear chunks of both strands loose such that there is no template to effect repairs.

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