Lecture 5 - Nucleic Acids, DNA Replication Flashcards
Nucleic Acids (4)
- Polymers specialized for storage and transmission of biological information.
- DNA: Deoxyribonucleic Acid
- RNA: Ribonucleic Acid
- Comprised of monomers called nucleotides.
Nucleotides have three components (3):
- nitrogen-containing base.
- pentose sugar
- phosphate group
Nucleotide vs. Nucleoside:
Nucleoside is a molecule consisting of just the nitrogen-containing base and pentose sugar (two components).
Nucleotide is a nucleoside with a phosphate group (three components).
Variations of Nitrogenous Bases:
Pyrimidines: single ring (T/U, C)
Purines: two fused rings (A, G)
Variations of Pentose Sugars:
- RNA contains the sugar RIBOSE which has additional -OH group (more polar)
- DNA contains DEOXYRIBOSE
Nucleotides can contain 1, 2, or 3 phosphates. Nucleotides that make up nucleic acids have _ phosphate group and are called…
1 phosphate group
nucleoside monophosphates.
Other roles for nucleotides (3)
- ATP, GTP: energy source in cellular work
- cAMP (cyclic AMP): essential for relaying extracellular cues to intracellular processes
- act as coenzymes in metabolic group transfer reactions (ex. FAD)
Formation of nucleic acids (3)
- phosphodiester linkage: formed when two nucleotides undergo condensation (anabolic) reaction.
- phosphate group of 5’ carbon of new nucleotide attaches to 3’ carbon of previous nucleotide. “grow in 5’-3’ direction”
- nucleic acids are also known as polynucleotides.
- oligonucleotides are short (~20 monomers)
Complementary base pairing is key to understanding ___ and ___ (4).
structure, function
- complementary base pairing: purines pair with pyrimidines by HYDROGEN bonds.
- DNA: A-T, G-C
- RNA: A-U, G-C
- A-T/A-U are double H-bonds, G-C are triple H-bonds and harder to break.
RNA structure (3)
- single-stranded
- base pairing can occur between different regions of the molecule, leading to diverse three-dimensional structures (structure is determined by order of bases)
- complementary base pairing can occur between RNA and DNA.
DNA structure (4 major points)
- double helix
- antiparallel strands
- bases exposed in major and minor groups
- right-handed
DNA structure: The Helix (2)
- sugar-phosphate backbone
- chains held together by…
- H-bonds between base pairs
- Van der Waals between adjacent bases on same strand.
DNA structure: Antiparallel strands (4)
- sugar-phosphate bonds determine strand direction.
- 5’ end has free 5’ phosphate group
- 3’ end has free 3’ -OH hydroxyl group
- 5’ end of one strand is paired with 3’ end of the other strand.
DNA structure: Base exposure (2)
- Minor groove: backbone of strands closer together.
- Minor groove: backbone of strands further apart.
DNA structure: Base exposure (4)
- Four possible configurations of base pairs in grooves
- Exposed base edges available for H-bonds with other molecules.
- Surfaces of A-T and G-C are chemically distinct.
- Proteins can recognize specific DNA sequences which is important for DNA replication and gene expression.
All DNA molecules have the same ____. Diversity is determined by ___.
structure, difference in nucleotide base sequences.
Why is DNA’s structure important? (4)
- stores genetic information: in the form of millions of nucleotides.
- susceptible to mutations: alterations in base sequences, genetic diversity.
- precisely replicated in cell division by complementary base pairing.
- genetic info is expressed as the phenotype: nucleotide sequence determines sequence of amino acids in proteins.
DNA transmits information in 2 ways:
- DNA can reproduce itself (replication)
- Certain DNA sequences can be copied into RNA (transcription). This RNA can specify sequence of amino acids in a polypeptide (translation)
Gene expression (defn)
is the process of transcription and translation.
DNA structure was determined by… (1)
- Physical evidence from X-ray cyrstallography.
- Biophysicist Maurice Wilkins discovered way to prepare DNA for X-ray diffraction.
- Rosalind Franklin analyzed DNA samples and found:
- DNA to be a double helix
- 10 nucleotides in each full turn (3.4 nm in length)- 2 nm diameter
DNA structure was determined by… (2)
- Chemical evidence from base composition.
- known that DNA is a polymer of nucleotides, and four different DNA nucleotides differed only by their bases.
- Erwin Chargraff reported that all DNA had the same about of purines and pyrimidines. CHARGAFF’S RULe (A=T, G=C)
Watson and Crick’s Model incorporated lines of evidence and published structure of DNA in 1953. (2)
- Antiparallel with sugar phosphate backbone.
2. Followed Chargaff’s rule, resulting in uniform width (OO=O, O=OO)
How is DNA replicated? (3)
- Semiconservative: each parent strand serves as a template and new molecule has one new and one old strand.
- Conservative: original DNA serves as template and original is not incorporated into new molecule.
- Dispersive: fragments of original DNA incorporated into new DNA.
The Meselson-Stahl Experiment (3)
- E-coli cultures grown with 15N then transferred to 14N
- Density gradient separation (centrifugation performed)
- Results explained semiconservative model. DNA is reproduced by semiconservative replication!
DNA replication requires 2 steps:
- Double helix is unwound, making two template strands.
2. New nucleotides form complementary base pairs with template DNA and are linked by phosphodiester bonds.
Formation of new DNA strands (3)
- free monomers that form DNA polymers have three phosphate groups.
- new nucleotides added to the growing 3’ end lose two phosphates (pyrophosphate) during phosphodiester bond formation.
- energy released by hydrolysis of dNTP drives condensation reaction.
6 Key players in DNA replication (6)
- DNA polymerase
- primase
- DNA helicase
- single-strand binding proteins
- sliding DNA clamp
- telomerase
Origins of replication (2)
- DNA replication begins with binding of the pre-replication complex to the origin of replication (ori)
- prokaryotes have 1
- eukaryotes have >1
- DNA is unwound and replication forks move away from one another.
DNA replication begins with a ___. (4)
Primer
- DNA polymerase requires a primer, a short “starter” strand - usually RNA
- Primase synthesizes a primer that is complementary to the DA template
- DNA polymerase adds nucleotides to the 3’ end of the primer
- Primer is degraded and DNA added in its place.
DNA polymerase (3)
- DNA polymerase substrates: dNTPS and template DNA
- “palm” brings active site and substrates together
- “fingers” recognize nucleotide bases
2 Proteins assist DNA polymerase
- DNA helicase uses energy from ATP hydrolysis to unwind DNA.
- Single-strand binding proteins keep the strands from reforming double helix.
Two types of newly replicating strands (2)
- Leading strand: grows continuously “forward” at its 3’ end as the form opens.
- Lagging strand: grows in shorter “backward” discontinuous stretches, known as Okazaki fragments.
- exposed 3’ end gets farther from the fork.
- each fragment requires own primer, synthesized by primase.
- segments linked by enzyme DNA ligase, resulting in a continuous strand of DNA.
DNA polymerases are fast because.. (2)
- they are processive: catalyze many linkages each time they bind to DNA.
- high processivity = greater number of nucleotides added before polymerase dissociates.
- polymerase-DNA complex is stabilized by a sliding DNA clamp - a protein that keeps the enzyme and DNA in close contact, increasing efficiency of polymerization.
In eukaryotes, DNA is ___ and the polymerase/replication complex is ____.
mobile, stationary.
- DNA goes in as one double stranded molecule, and emerges as two double stranded molecules.
Telomeres are not fully replicated (4)
- telomeres: repetitive sequences at the ends of eukaryotic chromosomes.
- in humans, sequence is TTAGGG, and repeated about 2,500 times.
- prevents DNA repair system from seeing chromosome end as a break.
- on lagging strand, terminal Okazaki primer is removed, no DNA can be synthesized to replace it.
- short piece of DNA removed.
- chromosome shortening with each replication; genes may be lost and cell dies.
Telomerase (2)
- continuously dividing cells have telomerase, which catalyzes addition of lost telomeres by using its own RNA sequence as a template (bone marrow stem cells and gamete producing cells)
- telomerase expressed in most cancer cells (anti-cancer target)
