Transcript DNA, RNA, and Protein Synthesis

Chapters 16 and 17
 Before the end of the semester we will be covering…
 Historical DNA experiments
 Structure of DNA/RNA
 DNA Replication
 Protein Synthesis (Transcription and Translation)
 Mutations
 Gene Expression (if time)
 No more labs this semester!
 Your final will be comprehensive  Both multiple
choice and short answer…more info to come!
Important Historical Experiments
 In the 1940s little was understood about inheritance and
how it worked.
 It was believed the genetic material was either DNA or
protein.
 It was understood that chromosomes are made of both DNA
and protein
 Initial experiments suggested it was protein…little was
understood about DNA’s structure or function (proteins
were identified as being more complex)
Avery, McLeod, McCarty
1944
 goal was to identify if inherited substance was
either DNA, RNA, or protein
 used chemicals and bacteria that only allowed
one of the above to be active at a time
 Results determined that the transforming agent
was DNA
 Scientific community still skeptical
Hershey and Chase 1952
 used bacteriophages to confirm DNA is the
genetic material
 bacteriophages were tagged with radioactive
isotopes (DNA--P; protein--S)
 it was shown that the DNA was able to “infect”
the bacteriophages, not the protein by
tracking the radioactivity
Watson and Crick
1953
 discovered the shape of DNA
molecule was a double helix
 using pictures of the molecule,
they built a model
 sugar/phosphate backbone
 nitrogen bases in the interior
 strands are antiparallel
 helix uniform in diameter
(base-pairing rules)
Wilkins and Franklin
1953
 Franklin used x-ray diffraction to
photograph DNA (her pictures were
used by Watson and Crick)
 Wilkins was working closely with
Franklin in her lab and allegedly showed
Watson and Crick the photograph that
helped them build their model
 Watson, Wilkins, and Crick were
awarded the Nobel Prize for science in
1962.
 Rosalind Franklin died in 1958 of cancer
and was never given a Nobel Prize.
 Using the original papers published in Nature in
1953, identify the characteristics of a DNA molecule.
 What important characteristics did they (Watson,
Wilkins, Crick, and Franklin) discover?
List/highlight as many as you can.
Deoxyribonucleic Acid (DNA)
 Classified as a nucleic acid
(biological molecule)
 Genetic material organisms
inherit from their parents
 Copied prior to cell division
(mitosis/meiosis)
 Shape of a double helix
 Made up of nucleotides
(building blocks)
 5-carbon sugar (deoxyribose),
phosphate, nitrogen base
 Base-Pairing Rules:
 AT
 CG
 Directionality
 Complementary strands
 Antiparallel: each strand runs in an
opposite direction

Designated 5’ and 3’ ends (carbon on
sugar)
 Two types of bases
 Purines: two carbon rings

Guanine, Adenine
 Pyrimidines: one carbon ring

Cytosine, Thymine, Uracil
Ribonucleic Acid (RNA)
 mRNA (messenger RNA)—instructions (from
DNA) for making protein
 tRNA (transfer RNA)– carries amino acids
 rRNA(ribosomal RNA)—makes up ribosomes
 5-carbon sugar (ribose)
 Single stranded (one gene)
 Uracil instead of thymine
 AU
Both RNA and DNA…
 Have adenine, cytosine,
and guanine
 Made of nucleotides
 Sugar and phosphate
backbone
 Nitrogen bases
perpendicular to
backbone
 Held together by
hydrogen bonds
Biochemical Gymnastics: DNA
Replication
 Occurs during S-phase of Interphase
in the cell cycle
 Semi-conservative process. Each
new strand of DNA produced is
made of one parental and one new
strand (described by Watson and
Crick)
 Each strand serves as a template for
the new strand
 In prokaryotes DNA is circular
 In eukaryotes DNA is linear
DNA Replication
(Overview)
 Begins at the origin of replication (specific sequences of
DNA nucleotides)
 Proteins recognize this sequence and attach to the DNA and
separate the two strands creating a “bubble”
 At either end of this “bubble” is the replication fork
 Replication then proceeds in both directions from the origin
until both strands are copied.
 In prokaryotes replication starts in one spot, in eukaryotes
multiple spots
The Players.
 Helicase: enzyme that unwinds and unzips the helix at the
replication forks.
 Single-strand Binding Protein (SSBP’s): binds to unpaired DNA
strand to keep them from re-pairing
 Topoisomerase: enzyme that relieves tension ahead of the
replication fork (from untwisting of strand)
 Primase: enzyme that synthesizes the RNA primer for replication
 DNA Polymerase: several enzymes that catalyze the synthesis of
new DNA (in eukaryotes there are 11 total); also checks for errors
 Ligase: links new fragmented DNA segments together
 Replication only occurs
in the 5’ to 3’ direction
 Nucleotides added only
to 3’ end of molecule…
 This is problematic for
one side of the DNA
molecule
 Leading strand:
continuous strand
 Single RNA primer
 Lagging strand:
discontinuous in
fragments (Okazaki
fragments)
 multiple RNA primers
The steps…
1. Origin of replication is located
2. Bubble forms in DNA helix by Helicase
3. Primase synthesizes primer to begin replication
 Need RNA primer to have something to add nucleotides to.
4. DNA Nucleotides are added by DNA polymerase to primer to
begin new strand
5. Replication proceeds in the 5’ to 3’ direction on both sides of
the molecule
 Leading and lagging strands
6. Replication continues until the entire molecule is copied
http://highered.mheducation.com/sites/0035456775/student_view0/chapter12/dna_replication.html
http://www.dnalc.org/resources/3d/04-mechanism-of-replication-advanced.html
Ending Replication…
 After every round of replication some of the DNA
molecule is lost due to polymerase not being able to
replicate it.
 To avoid excess loss of DNA, the ends of eukaryotic
chromosomes have telomeres (long repeating
sequences)


Excess DNA nucleotides (no genetic info)
Acts as a buffer to actual genes (does shorten over time—
thought to be evidence of aging)
Chapter 18
 Gene Expression: process by which
DNA directs the synthesis of proteins
 occurs in two parts: Transcription
and Translation

this process dictates the presence of
specific traits (genotype/phenotype)

occurs in all organisms
 Watson and Crick describes this as
the “central dogma”
(DNARNAProtein)
Transcription
 synthesis of RNA (mRNA) using DNA as a
template (protein instructions)
 occurs in nucleus (eukaryotes) or cytoplasm
(prokaryotes)

Prokaryotes can begin translation before
transcription is finished

Eukaryotes have an extra step during transcription
before translation can begin
 DNA is a template strand, which is used to produce
mRNA instructions (for protein)
 mRNA is complementary to DNA

uses different nucleotides (uracil)
 RNA polymerase unzips DNA and joins
complementary RNA nucleotides to copy
instructions
 reads 5’ to 3’ , no primer needed
 promoter: DNA sequence where RNA
polymerase attaches and initiates transcription
http://www.dnalc.org/resources/3d/13transcription-advanced.html
3 Stages
 Initiation
 RNA polymerase joins to the promoter and begins to
unwind helix
 helped by transcription factors (proteins), creates a
“transcription-initiation complex”
 Elongation
 RNA polymerase unwinds/untwist 10-20 nucleotides at a
time
 nucleotides added to 3’ end
 as mRNA is built, the molecule peels away from DNA and
the double helix reforms
 Termination
 in prokaryotes there is a terminator sequence (stop signal)
 eukaryotes transcribe a specific sequence to stop
transcription (creates pre-mRNA)
RNA Modifications
(eukaryotes only)
 RNA processing: enzymes in the nucleus modify pre-mRNA
 To help protect from degradation

5’ cap (modified G sequence)

3’ (poly-A tail)
 RNA splicing: removal of large portions of the RNA
molecule (cut and paste)
 eukaryotes have long stretches of non-coding DNA
interspersed with coding segments

introns: non-coding segments

exons: coding segments eventually expressed
RNA Splicing…
 introns are removed and
exons are joined together
 snRNPs: join together to
form a spliceosome
 Once finished,
completed mRNA leaves
nucleus to begin
translation
Translation
 synthesis of a polypeptide
using mRNA as
instructions
 occurs on ribosomes
(rRNA) in cytoplasm
 tRNA: transfers amino
acids to growing
protein

each associated with
a particular amino
acid

anticodon:
complementary RNA
sequence to mRNA
 instructions for producing a protein uses three
letters on mRNA (triplet code)
 codon: mRNA triplet (3 nucleotides)


methionine is “start”
“stop” codons UAA, UAG, UGA
3 stages
http://www.dnalc.org/resources/3d/16translation-advanced.html
 Initiation
 mRNA, tRNA, and ribosome start with amino acid methionine (initiator
tRNA)
 “translation initiation complex”
 Elongation
 amino acid added to chain via sites on ribosome (A-P-E)
 “elongation factors” help process; reads 5-3’
 requires energy
 Termination
 stop codons end synthesis, codes for “release factor”
 release factors bind and protein is released via hydrolysis
 Protein is then folded into appropriate shape with help of chaperonin
proteins
Oops!
http://www.bozemanscience.com/mutations/
 Mutation: change to genetic information
 Ultimate source of new genes
 May be spontaneous or result from mutagens
 Point mutations: change in single nucleotide (substitution)



silent: change doesn’t alter amino acid sequence
missense: changes one amino acid into another (minor
changes)
nonsense: change codon for amino acid into a stop codon
(premature end to translation)
 Frameshift mutation: add/lose nucleotides resulting in
change to reading frame of codons (not multiples of 3)
 Insertion or Deletion