Download DNA! - Chapter 10

DNA! - Chapter 10
Let’s review where we find
our genetic coding ……
What holds our genetic coding?
• Chromosomes
✓ Strands of DNA that contain all of
the genes an organism needs to
survive and reproduce
• Genes
✓ Segments of DNA that specify
how to build a protein
• There can be multiple genes
for a trait/protein
✓ Chromosome maps are used
to show the locus (location)
of genes on a chromosome
The E. Coli genome includes
approximately 4,000 genes
Genetic Variation
✓ Phenotypic variation among organisms is due to genotypic variation
(differences in the sequence of their DNA bases)
✓ Differences exist between species and within a species
• Different genes (genomes) → different proteins
• Different versions of the same gene (alleles)
• Differences in gene expression (epigenetics)
DO NOW:
What are the 3 components that
make up a nucleotide structure?
Structure of DNA
DNA stands for: Deoxyribonucleic acid
DNA is located in the nucleus of eukaryotic cells
and cytoplasm in prokaryotic cells
DNA and RNA are polymers composed of nucleotides
▪ Polymers = DNA and RNA
▪ Monomer = nucleotide
Each nucleotide contains a:
– Nitrogenous base
– 5-carbon sugar
– Phosphate group
Nitrogenous base
(A, G, C, or T)
Phosphate
group
Thymine (T)
Sugar
(deoxyribose)
The 4 Nitrogenous bases of DNA:
Thymine (T)
Cytosine (C)
Pyrimidines
Adenine (A)
Guanine (G)
Purines
What is the difference between
pyrimidines and purines?
Nitrogenous bases bond together to make a BASE
PAIR
Adenine → Thymine
How are
they bonded
Cytosine → Guanine
together?
How many
bonds does
each pair
have?
A sugar-phosphate
backbone is formed by
covalent bonding between
the phosphate of one
nucleotide and the sugar
of the next nucleotide
Nitrogenous bases extend
from the sugar-phosphate
backbone
What does Antiparallel mean?
● Two DNA strands run
parallel but in the
opposite alignment
● Allows nucleotides to
make the needed
hydrogen bonds
5’ - 3’
● These numbers identify the carbons on the sugar backbone
● Start to the right of the “o”
● Asymmetry gives a DNA strand “direction”
Count the carbons
Sugar-phosphate backbone
Phosphate group
Nitrogenous base
Sugar
DNA nucleotide
Phosphate
group
Nitrogenous base
(A, G, C, or T)
Thymine (T)
Sugar
(deoxyribose)
DNA nucleotide
DNA polynucleotide
Notice the →
● Different bonds
● Different bases
● 5’ - 3’ & 3’-5’
Sugar → Deoxyribose or Ribose
What is RNA?
●
●
●
●
Ribonucleic Acid, present in all living cells
Single Stranded
Helps with the creation of proteins!
Has 3 of the same nitrogenous bases DNA
has except uracil takes the place of thymine
DNA Replication
DNA Replication
• Cell Division (mitosis)
✓ Cells must copy their chromosomes
(S phase) before they divide so that each
daughter cell will have a copy
Questions before we start…
1. How does that DNA actually replicated?
2. What form is the DNA in during this?
AT
GC
CG
TA
GC
DNA Synthesis
✓ The DNA bases on each strand act
as a template to synthesize a
complementary strand
✓ The process is semiconservative
because each new double-stranded
DNA contains one old strand (template)
and one newly-synthesized
complementary strand
AT
GC
CG
ATTA
GGCC
CG
TA
GC
A
G
C
T
G
AT
GC
CG
TA
GC
T
C
G
A
C
Enzymes involved:
• Topoisomerase
✓ Unwinds (uncoils) the DNA
• Helicase
✓ An enzyme that separates strands of nucleic acids
“Replication Fork”
• Single Stranded Binding Proteins
✓ Prevent DNA that has been opened at the replication fork during DNA
replication from immediately reestablishing double helix conformation
• RNA Primase
✓ an enzyme that creates a short RNA sequence, called a
primer, tells the DNA polymerase where to start replication
• DNA Polymerase (I and III)
✓ Enzyme that catalyzes the covalent bond between the phosphate of one
nucleotide and the deoxyribose (sugar) of the next nucleotide
Polymerase I: replaces segments of
primer with DNA nucleotides
Polymerase III: binds at the end of the
primer and adds new DNA nucleotides
• Ligase
✓ Joins DNA strands back together (acts as a glue)
3’ end has a free deoxyribose
5’ end has a free phosphate
DNA polymerase:
✓ can only build the new strand in
the 5’ to 3’ direction
✓ Thus scans the template strand in
3’ to 5’ direction
DNA Replication Steps…
1. Initiation
2. Elongation
3. Termination
Initiation
• Primase (a type of RNA polymerase) builds an RNA primer
(5-10 ribonucleotides long)
• DNA polymerase attaches onto the 3’ end of the RNA primer
DNA
polymerase
Elongation
• DNA polymerase uses each strand as a template in the 3’ to 5’
direction to build a complementary strand in the 5’ to 3’ direction
• results in a leading strand and a lagging strand
DNA polymerase
Lagging Strand Creates Okazaki Fragments
• Newly synthesized DNA fragments that are formed on the lagging
template strand during DNA replication
Last step...
Termination
Primers are removed and
replaced with new DNA
nucleotides and the backbone is
sealed by DNA ligase
2 new daughter strands
of DNA!
Eukaryotic vs. Prokaryotic
DNA replication
Review of Enzymes involved
and DNA replication:
– DNA polymerase adds nucleotides to a growing chain
– DNA ligase joins small fragments into a continuous chain
– Helicase unwinds the DNA strand
– RNA Primase tells DNA polymerase where to start; “primer”
– Single Stranded Binding Proteins prevent DNA from coiling
back into a double helix during synthesis
Review...
Leading Strand
1. Topisomerase unwinds DNA and then Helicase breaks H-bonds
2. Primase creates a single RNA primer to start the replication
3. Polymerase slides along the leading strand in the 3’ to 5’ direction synthesizing
the matching strand in the 5’ to 3’ direction
4. The RNA primer is degraded by RNase H and replaced with DNA nucleotides by
DNA polymerase, and then DNA ligase connects the fragment at the start of the
new strand to the end of the new strand (in circular chromosomes)
Review...
Lagging Strand
1. Topisomerase unwinds DNA and then Helicase breaks H-bonds
2. Primase creates RNA primers in spaced intervals
3. Polymerase slides along the leading strand in the 3’ to 5’ direction synthesizing
the matching Okazaki fragments in the 5’ to 3’ direction
4. The RNA primers are degraded by RNase H and replaced with DNA nucleotides
by DNA polymerase
5. DNA ligase connects the Okazaki fragments to one another (covalently bonds the
phosphate in one nucleotide to the deoxyribose of the adjacent nucleotide)
Protein Synthesis
Protein Synthesis
• DNA provides the instructions for how to build proteins
• Each gene dictates how to build a single protein in prokaryotes
• The sequence of nucleotides (AGCT) in DNA dictate the order
of amino acids that make up a protein
Nucleotide sequence of His gene
Protein Synthesis
• Protein synthesis occurs in two primary steps
1
mRNA (messenger RNA)
copy of a gene is
synthesized
✓Cytoplasm of prokaryotes
✓Nucleus of eukaryotes
2
mRNA is used by ribosome to
build protein
(Ribosomes attach to the
mRNA and use its sequence of
nucleotides to determine the order
of amino acids in the protein)
✓Cytoplasm of prokaryotes
and eukaryotes
✓Some proteins feed directly into
rough ER in eukaryotes
Protein Synthesis
• Transcription
Initiation
1) INITIATION
✓ RNA polymerase binds to a
region on DNA known as the
promoter, which signals the
start of a gene
✓ Promoters are specific to genes
✓ RNA polymerase does not need
a primer
✓ Transcription factors assemble
at the promoter forming a
transcription initiation complex
– activator proteins help stabilize
the complex
✓ Gene expression can be regulated (turned
on/off or up/down) by controlling the amount
of each transcription factor
(eukaryotes)
Protein Synthesis
1) INITIATION
• Transcription
Elongation
✓ RNA polymerase unwinds
the DNA and breaks the
H-bonds between the bases
of the two strands, separating
them from one another
✓ Base pairing occurs between
incoming RNA nucleotides
and the DNA nucleotides of
the gene (template)
• recall RNA uses uracil
instead of thymine
AGTCAT
UCAGUA
Protein Synthesis
• Transcription
Elongation
✓ RNA polymerase unwinds
the DNA and breaks the
H-bonds between the bases
of the two strands, separating
them from one another.
✓ Base pairing occurs between
incoming RNA nucleotides
and the DNA nucleotides of
the gene (template)
• recall RNA uses uracil
instead of thymine
5’
3’
+ ATP
5’
✓ RNA polymerase catalyzes bond to
form between ribose of 3’ nucleotide
of mRNA and phosphate of incoming
RNA nucleotide
3’
+ ADP
Protein Synthesis
• Transcription
Elongation
The gene occurs on only one of the DNA
strands; each strand possesses a separate
set of genes
Protein Synthesis
1) INITIATION
• Transcription
Termination
✓ A region on DNA known as
the terminator signals the
stop of a gene
✓ RNA polymerase disengages
the mRNA and the DNA
Protein Synthesis
• Alternative Splicing (eukaryotes only)
✓ Exons are
“coding” regions
✓ Introns are removed
✓ different combinations
of exons form
different mRNA
resulting in multiple
proteins from the
same gene
✓ Humans have 30,000
genes but are capable
of producing 100,000
proteins
Protein Synthesis
Transcription
tRNA
synthesis
1
2
mRNA
mRNAmRNA copy of a gene
is synthesized
✓Cytoplasm of prokaryotes
✓Nucleus of eukaryotes
mRNA is used by ribosome to
build protein
(Ribosomes attach to the
mRNA and use its sequence of
nucleotides to determine the order
of amino acids in the protein)
✓Cytoplasm of prokaryotes
and eukaryotes
✓Some proteins feed directly into
rough ER in eukaryotes
Translation
Protein Synthesis
Transcription
• Translation
tRNA
synthesis
✓ Every three mRNA nucleotides (codon) specify an amino acid
mRNA
Translation
Protein Synthesis
• Translation
✓ tRNA have an anticodon region that specifically binds to its codon
Protein Synthesis
Transcription
• Translation
✓ Each tRNA carries
specific amino acid
tRNA
asynthesis
mRNA
Translation
Protein Synthesis
Transcription
tRNA
synthesis
mRNA
Translation
Aminoacyl tRNA synthetases attach
amino acids to their specific tRNA
Protein Synthesis
mRNA
• Translation
Initiation
✓ Start codon signals where the gene
begins (at 5’ end of mRNA)
Translation
5’
3’
AUGGACAUUGAACCG…
start codon
Protein Synthesis
Small ribosomal subunit
• Translation
Initiation
✓ Start codon signals where the gene
begins (at 5’ end of mRNA)
✓ Ribosome binding site (Shine
Dalgarno sequence) upstream from
the start codon binds to small
ribosomal subunit
– then this complex recruits the
large ribosomal subunit
Small ribosomal subunit
Large ribosomal subunit
Ribosome
Protein Synthesis
• Translation
Scanning
✓ The ribosome moves in 5’ to 3’ direction “reading” the mRNA and
assembling amino acids into the correct protein
large ribosome subunit
small
ribosome
subunit
Protein Synthesis
• Translation
Scanning
✓ The ribosome moves in 5’ to 3’ direction “reading” the mRNA and
assembling amino acids into the correct protein
Protein Synthesis
• Translation
Termination
✓ Ribosome disengages from the mRNA
when it encounters a stop codon
Practice Question
Translate the following mRNA sequence
AGCUACCAUACGCACCCGAGUUCUUCAAGC
Practice Question
Translate the following mRNA sequence
AGCUACCAUACGCACCCGAGUUCUUCAAGC
Serine – Tyrosine – Histidine – Threonine – Histidine – Proline – Serine – Serine – Serine - Serine
Practice Question
Translate the following mRNA sequence
AGCUACCAUACGCACCCGAGUUCUUCAAGC
Serine – Tyrosine – Histidine – Threonine – Histidine – Proline – Serine – Serine – Serine - Serine
Ser – Tyr – His – Thr – His – Pro – Ser – Ser – Ser - Ser
Practice Question
Translate the following mRNA sequence
AGCUACCAUACGCACCCGAGUUCUUCAAGC
Serine – Tyrosine – Histidine – Threonine – Histidine – Proline – Serine – Serine – Serine - Serine
Ser – Tyr – His – Thr – His – Pro – Ser – Ser – Ser - Ser
S – Y –H– T – H – P – S – S – S - S
Protein Synthesis
• Multiple RNA polymerases can
engage a gene at one time
• Multiple ribosomes can engage
a single mRNA at one time
Transcription
DNA
mRNAs
Translation
Protein Synthesis
• Eukaryotes:
transcription occurs
in the nucleus and
translation occurs in
the cytoplasm
• Prokaryotes:
Transcription and
translation occur
simultaneously in
the cytoplasm
RNA
• There are four main types of RNA:
1. mRNA
- RNA copy of a gene used as a template for protein synthesis
2. rRNA
- part of structure of ribosomes
3. tRNA
- amino acid carrier that matches to mRNA codon
4. snRNA
- found in nucleus where they have several important jobs
Practice Questions
1. Why is DNA synthesis said to be “semiconservative”?
2. What role do DNA polymerase, DNA primase (a type of RNA polymerase),
helicase, topoisomerase, RNase H, and ligase play in DNA replication?
3. What is the difference between how the leading strand and lagging strand are
copied during DNA replication? Why do they have to be synthesized differently
in this fashion?
4. What would happen if insufficient RNase H were produced by a cell? What if
insufficient ligase were produced by a cell?
5. What are four key differences between DNA polymerase and RNA polymerase?
(“they are difference molecules” doesn’t count as one!)
6. Compare and contrast codons and anticodons?
7. What is alternative splicing? Why is it necessary in eukaryotes?
8. During translation, what amino acid sequence would the following mRNA
segment be converted into: AUGGACAUUGAACCG?
9. How come there are only 20 amino acids when there are 64 different codons?
10. How come prokaryotes can both transcribe and translate a gene at the same time,
but eukaryotes cannot?