DNA Replication Flashcards
Which nitrogenous bases are pyrimidines and purines?
adenine and guanine are purines
thymine and cytosine are pyrimidines
What happens in the INITIATION stage of DNA replication?
- DNA helicase enzyme unwinds the double helix by breaking the hydrogen bonds (starts at a specific nucleotide sequence called the origin of replication; a bubble and replication fork is formed)
- DNA gyrase (topoisomerase) enzyme assists by making sure twists and knots don’t form ahead of the bubble (this helps relieve strain at the replication fork)
- Single-stranded binding proteins (SSBs) keep DNA template strands separated
What is DNA packaged around? What are these structures called in eukaryotic and prokaryotic cells?
packing around protein complexes known as HISTONES
called a nucleosome in eukaryotic cells; called a nucleoid in prokaryotic cells
When does the helix make one turn? What happens because of this?
makes one turn every 10 nucleotides or 3.4 nm; therefore there is 0.34 nm between base pairs
What does DNA polymerase I do?
replaces the RNA primer with DNA, leaving a tiny gap
What is the Chargaff’s Rule?
the proportion of A always equals the proportion of T AND the proportion of C always equals the proportion of G
A+G = T+C [ A=T and C=G]
What is semiconservative replication? who was it proposed by and confirmed by?
each new molecule of DNA contains 1 strand of the original parent DNA & one new complementary daughter strand (hybrid model)
~ each new DNA conserves half of the original molecule
proposed by Watson & Crick; confirmed by Meselson & Stahl
What are genes? What do they do
the function units of DNA; they sequence codes for the production of specific proteins or RNA
How many antiparallel strands of nucleotides does DNA consist? How do they work
consists two
one strand runs from 5’ to 3’ and the other runs from 3’ to 5’
~ the 3’ end terminates with a hydroxyl group of the deoxyribose sugar
~ the 5’ end terminates with the phosphate group
what are purines and pyrimidines? What do they pair up with?
purines are double-ring structures (adenine and guanine) and pyrimidines are single-ring structures (thymine and cytosine)
purines pair up with pyrimidines through hydrogen bonding
what is DNA replication
the process of producing two identical DNA molecules from an original parent DNA molecule
What is the lagging strand?
synthesized away from the replication fork; formed discontinuously (in segments) from 5’ end
Why does telomerase activity slow down over time?
- in early childhood, telomerase, which replaces the telomeric region, is quite active
- with aging, the enzyme activity slows and genetic info can be lost; chromosomes can lose coding portions of their DNA as the tips fray and get damaged
What are single stranded binding proteins (SSBs)?
proteins that keep DNA template strands separated
What are okazaki fragments? what are they made on?
short DNA segments, each beginning with an RNA primer; made on lagging strand
What is the leading strand?
synthesized towards the replication fork; formed continuously from the 3’ end of the parent strand
What is DNA polymerase III?
the enzyme that catalyzes the addition of new nucleotides
What are the nucleotides in DNA supported by?
a deoxyribose sugar and a phosphate backbone
What did Rosalind Franklin discover? What year was she active?
- used a technique called X-ray diffraction to analyze the structure of biological molecules (making DNA interact with X-rays which are bent by the molecules they encounter)
- discovered that DNA has a helical structure with two regularly repeating patterns
- stated that nitrogenous basses (ATCG) were located on the inside of the helical structure and the sugar-phosphate backbone was located on the outside
active in 1950s
What is primase
enzyme that creates an RNA primer which terminates with a 3’ hydroxyl group, allowing the DNA polymerase III to start attaching new nucleotides
What is the origin of replication
a specific nucleotide sequence where replication is initiated
How many DNA polymerases are in prokaryotes and eukaryotes?
PROKARYOTES: 5
EUKARYOTES: 13
What is the rate of DNA replication in eukaryotes and prokaryotes? Compare them.
PROKARYOTES: faster (about 1000 nucleotides per sec)
EUKARYOTES: less than prokaryotes due to more elaborate enzyme complexes required in replication and a strict proofreading mechanism (40 nucleotides added per second)
What is DNA polymerase II?
catalyzes the repair of nucleotide base pairs & proofreads newly synthesized DNA
What did Francis Crick and James Watson discover? When were they active?
- emphasized that DNA has a twisted, ladder-like structure called a double helix
~ the sugar-phosphate molecules make up the sides of the ladder, and the bases make up the rungs by projecting inward at regular intervals along each strand - determined that the distance between the sugar-phosphate handrails remained constant over the length of the molecule
~ however adenine and guanine have a double-ring structure & thymine and cytosine have a single-ring structure
DISCOVERED THE MOLECULAR STRUCTURE OF DNA
active in 1950s
Do both eukaryotes and prokaryotes have primers? telomeres?
primers - both have
telomeres - eukaryotes have; prokaryotes don’t
How are errors with DNA replication dealt with? What’s the error rate?
when nucleotides are being added into the new strand, sometimes the wrong one is added
~ DNA polymerase III can reverse and replace the nucleotide during replication
~ DNA polymerase II (and other enzymes) can replace the nucleotide after, fixing the mismatch (also used to fix damage due to environmental factors)
error rate is about one per billion nucleotide pairs
What is the TERMINATION step of DNA replication?
- each of the two new strands begins to twist and coil into their double helix form
- when the new DNA strands are completed, they separate from each other
- the replication machine (protein-DNA complex at each form) is dismantled
How many origins of replication are in prokaryotes and eukaryotes?
PROKARYOTES: smaller circular chromosome contains A SINGLE origin of replication
EUKARYOTES: larger linear chromosome may contain THOUSANDS of origins of replication
What is topoisomerase/gyrase?
enyzme that assists by making sure knots and twists don’t form ahead of the bubble
What is DNA ligase?
enzyme that joins together backbone gaps
What are telomeres?
a repetitive section of DNA near each end of a chromosome that helps protect against the loss of important genetic information during replication
~ shortens with every round of replication
What did Fredrich Miescher discover? What year was he active?
- isolated nuclei of white blood cells and from these nuclei, he extracted a weakly acidic substance containing nitrogen and phosphorus which he called NUCLEIN; this was later known as NUCLEIC ACID
- discovered DNA
active in 1869s (pre 1920s)
How many hydrogen bonds do A-T and C-G have?
A-T have 2 hydrogen bonds
C-G have 3 hydrogen bonds
what is the direction of DNA replication
5’ to 3’ direction
Who proposed that the double helix turns in a clockwise (or right handed) direction?
James Watson and Francis Crick
what is DNA helicase?
enzyme that unwinds the double helix by breaking the hydrogen bonds ; starts at the origin of replication
What are the three stages of DNA replication? Give a description of each.
- Initiation
~ helix is unwound to expose bases for new base pairing - Elongation
~ new strands are assembled using parental DNA as template - Termination
~ process stops and new DNA molecules separate
What happens in the DNA ELONGATION step of DNA replication?
- new nucleotides are joined together, one at a time to create a strand of DNA that is complementary to the parental strand
- when DNA is unwound, there is no free 3’ hydroxyl end on either single strand, so the DNA has to be “primed”; primase enzyme creates an RNA primer which terminates with a 3’ hydroxyl group, allowing the DNA polymerase III to start attaching new nucleotides
- DNA polymerase III then builds the complementary strand by attaching nucleotides to the 3’ end and previously placed nucleotides
- once DNA polymerase III reaches an ‘old upstream primer, DNA polymerase I replaces the RNA with DNA leaving a tiny gap (gaps are joined together by DNA ligase)
What did Frederick Griffith discover? What year was he active?
- executed a experiment that involved two strains of the bacterium Streptococcus pneumoniae (one being lethal to mice and the other being harmless)
- the S strain type was highly pathogenic but could be made non-pathogenic by heating it; R strain was non-pathogenic
- discovered that mice died when injected with a mixture of heat-killed S-strain bacteria and living R-strain bacteria
- concluded that S-strain had somehow passed on its deadly properties to the live, non-pathogenic R-strain
~ called this the TRANSFORMING PRINCIPLE as something from the heat-killed pathogenic bacteria must have transformed the living non-pathogenic bacteria to make them deadly
active in 1928
What did Oswald Avery do? What year was he active?
- supported Frederick’s study on how DNA was the hereditary material
- prepared identical extracts of the heat-killed S Strain by growing the cells in liquid culture, isolating the bacteria, disrupting the cell membranes, and collecting the cell contents
- one of 3 enzymes was added to each extract (one destroyed protein, one destroyed RNA, and one destroyed DNA)
- the enzyme-treated extract was then mixed with the live R-strain cells; the one that did not cause any transformation of the R-strain to the pathogenic S-strain was the extract with the DNA-destroying enzyme
- this showed that DNA caused the transformation; the transforming principle was DNA
active in 1944
What did Francis Collins discover? What year was he active?
- researched the genome for genes that cause human disease
- pinpointed the gene that causes cystic fibrosis, Duchenne muscular dystrophy, and Huntington’s disease
active in 1989
What did Martha Chase and Alfred Hershey discover? What year were they active?
- executed a blender experiment that determined that phage DNA was the genetic material and not protein
- used the T2 bacteriophage strain of virus which has a protein coat that surrounds DNA; when infecting a bacterial cell, the virus injects its genetic information into it
~ the remaining viral structure stays attached to the outside of the bacterium and is called a bacteriophage “ghost” - a radioactively labelled Dna was allowed to infect E. colli bacteria
- aimed to separate the bacteriophage ghosts and a pellet of bacteria; they discovered that most of the radioactively labelled protein was in the liquid medium and not in the bacteria
~ this demonstrates how protein is not a hereditary material as it was a part of the bacteriophage ghosts instead of injecting itself into the bacterial cells
active in 1952 (1950-1954)
What are some similarities that prokaryotes and eukaryotes have in terms of DNA replication?
- both require origins of replication
- have elongation occur in 5’ to 3’ direction
- have continuous synthesis of a leading strand and a discontinuous synthesis of a lagging strand
- require primers to synthesize okazaki fragments
- use DNA polymerase enzymes
What did Craig Venter do?
- using a new technique, called “expressed sequence tags (ESTs)”, he sequenced the human genome and identified thousands of human genes
- he and his team created the first synthetic organism, Mycoplasma mycoides JCVI-syn1.0, by synthesizing the entire genome of a bacterium, inserting it into a cell, and demonstrating that the cell could function and replicate with the synthetic genome
What did Matthew Meselson and Franklin Stahl discover? When were they active?
- discovered that DNA replication was semi-conservative
- grew E, Colli bacteria in a medium that contained a heavy isotope of nitrogen (15N)
- transferred the bacteria to a culture medium that had 14N as a source of nitrogen; samples of cells and their DNA were removed before this transfer and isolated
- noticed that DNA samples taken before the transfer had the same density and appeared as a distinct band which meant that DNA had 15N
- after 1 round of replication, the sample formed a single band and its position indicated that its density was midway between both nitrogens;
~ concluded that DNA was composed of 1 strand labelled with 14N and 1 strand labelled with 15N - after the 2nd round of replication, the DNA sample separated into two distinct bands after centrifugation; one band corresponded to 14N DNA only and the other corresponded to DNA that had 1 strand labelled with 14N and the other with 15N
active in 1950s
