Ch 8 Flashcards
Describe how DNA serves as genetic info
the structure of DNA helps explain two primary features of biological info storage.
-first, the linear sequence of bases provides the actual info
-genetic info is encoded by the sequence of bases along the strand of DNA, in much the same way as our written language uses linear sequence of letters to form words and sequences
Describe and recognize the process of DNA replication (aka DNA synthesis), including the function of the necessary enzymes and directionality of replication. Be able to repeat the replication fork
DNA synthesis begins at the origin of replication (modeled
as oriC) in prokaryotes
Formation of a replication fork must occur: zone of
unwound DNA where DNA polymerization is occurring
steps in DNA synthesis (DNA replication)
- DNA topoisomerases like DNA gyrase
remove the supercoils in the DNA to create linear DNA
steps in DNA synthesis (DNA replication)
- DNA topoisomerases like DNA gyrase
remove the supercoils in the DNA to create linear DNA
steps in DNA synthesis (DNA replication)
- DNA is nicked: DNaA proteins bind to the
origin and cleave the hydrogen bonds between complementary strands
steps in DNA synthesis (DNA replication)
- Next, DNA helicase acts to unwind
the DNA for a short distance and expose it as 2 single-stranded regions
steps in DNA synthesis (DNA replication)
- next, single-stranded binding proteins (SSBs) bind to these exposed
regions and prevent them from re-annealing to one another
steps in DNA synthesis (DNA replication)
- an RNA primer is synthesized against the template
ssDNA by primase
steps in DNA synthesis (DNA replication)
- Extension of DNA or polymerization (adding 3’ end) is performed by DNA polymerase III
occurs continuously on the leading strand
occurs discontinuously on the lagging strand creating okazaki fragments
new primers have to be added as
the fork ‘moves’ and exposes new
DNA ahead of the primer
steps in DNA synthesis (DNA replication)
- Once DNA synthesis is complete along the template the RNA primers must be
removed and replaced by DNA nucleotides
the RNA primers are removed by
DNA polymerase I proofreading
after the synthesis step
steps in DNA synthesis (DNA replication)
- Okazaki fragments are sealed together by DNA ligase to form
a continuous piece of DNA (no RNA nucleotides)
semi-conservative DNA replication
each chromosome now has 1 single strand of parental DNA (template) and one newly synthesized strand of DNA (daughter)
major enzymes in DNA synthesis
helicase
unzipping the DNA helix
major enzymes in DNA synthesis
gyrase
helping to untangle the DNA supercoils
major enzymes in DNA synthesis
primase
synthesizing an RNA primer
major enzymes in DNA synthesis
DNA polymerase III
adding bases to the new DNA chain; proofreading the chain for mistakes
major enzymes in DNA synthesis
DNA polymerase I
removing primer, closing gaps, repairing mismatches
major enzymes in DNA synthesis
ligase
final bonding of nicks in DNA during synthesis and repair
Describe and recognize the differences between DNA and RNA.
DNA
- deoxyribose sugar
- double stranded
-bases: Adenine-Thymine Cytosine-Guanine
RNA
-ribose sugar
-single stranded
-bases: Adenine-Uracil Cytosine-Guanine
Transcription Steps
- transcription begins when RNA polymerase bind to the
promoter sequence
Transcription Steps
- transcription proceeds in the
5’->3’ direction
Transcription Steps
- transcription stops when it reaches the DNA-based
terminator sequence
Transcription Steps
- the mRNA transcript looks just like the coding strand (U’s for T’s) because
RNA polymerase adds nucleotides complementary to the template strand (5’ to 3’)
What enzyme performs transcription?
RNA polymerase
Steps in Translation: Initiation
30S and 50S ribosomal subunits assemble with
mRNA and Met-tRNA
Steps in Translation: Initiation
- process is aided by several proteins called
initiation factors (IF’s)
Steps in Translation: Initiation
- begins when the AUG start codon is situated in the
P site (peptidyl site) of the ribosome
Steps in Translation: Initiation
- allows the anticodon portion of Met-tRNA to bind to the
AUG of the mRNA
Steps in Translation: Elongation
amino acids are brought into the ribosome to
elongate the polypeptide, then the ribosome is translocated to a new codon
Steps in Translation: Elongation
- A (aka receptor) site is filled:
Aminoacyl tRNAs (tRNA + amino acid) are brought to the A site of the ribosome (specific amino acid selected by complementary tRNA anticodon- mRNA codon binding)
this is aided by protein elongation factors (EFs)
Steps in Translation: Elongation
- Transpeptidation
the amino acids in the P site join the new amino acid in the A site via formation of a peptide bond between aa’s
Steps in Translation: Elongation
- Translocation
movement of the ribosome along the mRNA occurs to move a new codon into the A site
Steps in Translation: Elongation
- Translocation
how does it work?
A site –> P site
P site –> E site (aka exit)
A site will be emptied so a new aminoacyl tRNA can enter it
uncharged tRNA in the E site dissociates from the ribosome completely
Steps in Translation: Termination
occurs when a stop codon enters the
ribosome’s A site
aided by protein release factors (RF’s) which recognize stop codon
Steps in Translation: Termination
- completed polypeptide is cleaved from tRNA and ribosome subunits
dissociate
Steps in Translation: Termination
- subunits are free to form new
initation complexes and repeat the entire process on a new mRNA strand
coupled transcription/translation,
Prokaryotes are much more
efficient at protein expression
than eukaryotes
- Since there is no membrane to
separate the DNA from the
cytoplasm, translation of
mRNA can start as soon as a
transcript is produced
transpeptidation
the amino acids in
the P site join the new amino acid in
the A site via formation of a peptide
bond between aa’s
translocation
movement of the
ribosome along the mRNA occurs to
move a new codon into the A site
mutation:
change in genetic material- within the nucleotide sequence
point mutation:
a single base (nucleotide) change
only affects 1 codon
missense mutation
results in a different amino
silent (aka neutral) mutation
results in the same amino acid code still being encoded
nonsense mutation
results in the conversion of an affected codon from endoding an amino acid to becoming a stop or nonsense codon
frameshift mutation
results in multiple changes
transformation
Naked (free) DNA/plasmids in
the environment can be taken
up by a cell
- For successful HGT &
recombination: this DNA must
recombine into chromosome
by homologous recombination - Exception: a whole plasmid can
be taken up and remain free in
the cytoplasm! - Non-homologous genes will be
degraded (= unsuccessful
HGT/recombination)
transduction
HGT mediated by a bacteriophage
(bacterial virus) carrying DNA into a host as it
‘infects’ the host
- A incorrectly packaged bacteriophage carries bacterial
chromosomal DNA into the cell it infects - Called a transducing phage
- Cell can gain new genes via homologous recombination of
the introduced DNA (if HGT is successful) - If there is no homologous genes in the new host, the
transferred DNA could also be degraded (this would be
unsuccessful HGT)
transposons
Segments of DNA that can move from one region of DNA to
another = ‘jumping genes’
- Contain a gene for the transposase, enzyme that helps it move
(for cutting and resealing DNA) - Insertion sequences only have the transposase gene
- Composite (complex) transposons carry other genes plus
transposase (which is often sandwiched by two insertion
sequence
conjugation
Cell-to-cell transfer of a plasmid. The original cell retains the
plasmid and one strand moves into the recipient cell (where
the other strand is replicated)
- Also called mating but its not sexual reproduction!
Requirements for conjugation:
* Cell-to-cell contact between donor & recipient cells via a
pilus made by the donor cell
- tra genes - these encode the pilus and other apparatus
necessary for conjugation - oriT (origin of transfer): this is the site where rolling-circle
replication begins to make a copy of the plasmid DNA that
is exchanged with the neighboring cell (in contact via pilus)
