17- gene to protein.pdf

How does DNA control a cell?
◦ By controlling Protein Synthesis.
◦ Proteins are the link between
genotype and phenotype.

What makes a firefly glow?

Central Dogma of Gene
Expression

Complete the Comparison chart for DNA and RNA

DNA and RNA are both Nucleic Acids
Structure DNA RNA
Sugar
present
Nitrogen
bases
present
Number of
Strands
Relative
length
deoxyribose
ribose
A, C, G, T A, C, G, U
2 1
Long, contains
hundreds or
thousands of
genes
1 3
types one three
Shorter, about the
length of one gene
SIMPLE
SKETCH
mRNA
rRNA
tRNA

The Genetic Code
◦ Sequence of bases in DNA correspond
to the sequence of amino acids in
proteins.
◦ There are only 4 bases but 20 naturally
occurring Amino Acids.
◦ “Codon” = three consecutive bases
code for each amino acid
◦ One gene codes for one protein

Codon Dictionary
◦ Start- AUG (Met)
◦ Stop- UAA
UAG
UGA
◦ 60 codons for the
other 19 AAs.

Dictionary of the genetic code
◦ Each amino acid
except “start” has
multiple codons.
◦ This helps safeguard
against error in
transcription (writing)
or translation
(reading) the code.

Code Redundancy
◦ Third base in a codon shows
"wobble”.
◦ First two bases are the most important
in reading the code and giving the
correct AA.
◦ The third base often doesn’t matter.

Comment
◦ "Wobble" effect allows for 45 types
of tRNA instead of 61.
◦ Reason - in the third position, U can
pair with A or G.
◦ Inosine (I), a modified base in the
third position can pair with U, C, or
A.

Importance of Wobble
◦ Allows for fewer types of tRNA.
◦ Allows some mistakes to code for
the same AA which gives exactly
the same polypeptide.

Reading Frame
◦ The “reading” of the code is every three
bases.
◦ Ex: the red cat ate the rat
◦ Ex: ATT GAT TAC ATT
◦ The “words” only make sense if “read”
in this grouping of three.

Code Evolution
◦ The genetic code is nearly universal.
◦ Ex: CCG = proline (all life)
◦ Reason - The code must have evolved
very early. Life on earth must share a
common ancestor.
◦ Biotech applications use the universal
nature of DNA to move genes from
species to species.

Flow of
genetic info
◦ Transcription - to
write the code to
mRNA
◦ Site of transcription
is the cytosol
(bacteria) or nucleus
◦ Translation - to read
and relate the code
with tRNA and rRNA
◦ Site of translation is
the ribosome

Transcription – the DNA directed
synthesis of RNA
◦ Promoter – specific
nucleotide sequence
along the DNA
◦ In Eukaryotes these
promoters include a
TATA box
◦ Terminator – end of
transcription = UAA,
UAG & UGA

Step by Step through Transcription
◦ INITIATION
◦ RNA polymerase
binds to promoter
◦ This cause the DNA
to unwind & separate
◦ RNA transcript
begins to form

ELONGATION
The template strand is
transcribed by adding
complementary RNA
nucleotides in 5’ 3’
direction
◦ The RNA transcript
grows and DNA
reforms double helix
◦ In Prokaryotes, the
transcript is
immediately usable as
mRNA

TERMINATION
◦ Once a termination
sequence is
transcribed, the
RNA transcript is
released and RNA
polymerase
detaches from DNA
◦ There are three
termination codes

RNA Modification in Eukaryotic cells
◦ 5’ cap and Poly- A tail are added to RNA
transcript
◦ Prevents breakdown of DNA
◦ Facilitates passage through nuclear
pores
◦ Cap provides point of attachment at
ribosome

RNA Processing - RNA Splicing
◦ INTRONS – non-coding segments of
eukaryotic genes
◦ EXONS – coding segments
◦ During RNA processing, introns are
removed and exons are spliced together

RNA Processing - RNA Splicing
◦ Spliceosomes – an
assembly of
snRNPS -
(ribonuclear
proteins) + other
proteins bind to the
end of introns –
recognize code
◦ Newly spliced exon
–only mRNA can
exit nucleus

Functional and Evolutionary
importance of Introns
◦ Introns may play
regulatory roles in the
cell.
◦ Benefit of split genes is
to enable a single gene
to encode more than
one kind of protein.
◦ Facilitates the shuffling
among genes
promoting evolution

Alternative Splicing
◦ The RNA can be spliced into different
mRNA’s.
◦ Each different mRNA produces a
different polypeptide.
◦ Ex. – this explains how the body can
create infinite number of antibodies.

Another Example
◦ Bcl-X L – inhibits apoptosis
◦ Bcl-X S – induces apoptosis
◦ Two different and opposite effects!!

DSCAM Gene
◦ Found in fruit flies
◦ Has 100 potential splicing sites.
◦ Could produce 38,000 different
polypeptides
◦ Many of these polypeptides have been
found

Commentary
◦ About 60% of genes are estimated
to have alternative splicing sites.
◦ One gene does not equal one
polypeptide.
◦ The work of Beadle & Tatum is
being undone with the power of
technology and better
experimentation!

Transcription Review
◦ A biologist inserts a gene from a human
liver cell into a bacterial chromosome. The
bacterium transcribes the gene into mRNA
and translates it immediately into a protein.
The protein produced is useless and is
found to have many extra amino acids than
does the protein made by the eukaryotic
liver cell. Explain Why.

Protein Synthesis
◦ Translation is the
RNA-directed
synthesis of a protein.
◦ one codon one
amino acid
◦ Transfer RNA (tRNA)
molecules interpret
the genetic code as
written on the mRNA
transcript

Role of Transfer RNA
◦ Links mRNA codon
to its matching
amino acid.
◦ “Anticodon” –
complementary
base sequence to
mRNA codon.
◦ Amino Acid
attachment site is
located opposite
the anticodon.

Ribosomes
◦ Structure - Large and
small subunit both
composed of protein
and rRNA.
◦ One enzyme for each
amino acid
◦ Facilitate
• codon – anticodon
complex formation
• peptide bond
formation
◦ Prokaryotic ribosomes
are often the target of
antibiotics

Recognition steps for accurate
translation
◦ 1. Correct match between the tRNA
and the amino acid
◦ Each amino acid has a specific enzyme
that aids in attachment to tRNA –
aminoacyl-tRNA synthetase
◦ 2. Codon-anticodon bonding insures
translation

Initiation of Translation
◦ mRNA binds to a small ribosomal subunit –
AUG is the “start” sequence.
◦ GTP provides energy needed to bring the
large subunit to create a complex.
◦ E = exit ; P = protein building site and A =
enzyme driven Amino Acid binding site.

Elongation of polypeptide chain stop
◦ 1. Codon
recognition
◦ 2. rRNA serves as
a ribozyme –
catalyst of peptide
bond
◦ 3. translocation –
ribosome shifts
the mRNA by one
codon – both
move relative to
eachother

Termination
◦ Stop codon reaches “A” site.
◦ Release factor matches stop codon &
protein is released.
◦ Ribosome complex disintegrates.

Free vs. Bound Ribosomes
◦ Free Ribosomes
• Protein is needed in cytosol
◦ Bound ribosomes
• Protein is needed at membrane or
beyond via ER
• True of cells that line secretory organs
or tissues

Signal recognition particle carries the
complex to the ER membrane
20 amino
acids long
◦ Signal recognition protein temporarily binds
ribosome to the ER membrane
◦ Protein is fed into cisterna as it elongates

Mutations –changes in DNA
◦ Chromosomal mutations - large scale
changes to chromosomes
◦ Point mutations – small scale changes
in just one base pair of a gene
• Substitution
• Insertion
• Deletion
Drosophila antennapedia gene
mutation

Mutations
◦ If Mutation occurs in a gamete or cell
that creates gametes, it may be
transmitted.
◦ If mutation has an adverse affect on
phenotype it is called a genetic disorder
or hereditary disease.

Molecular Basis of Sickle-cell disease
◦ One change in the base sequence leads
to the production of an abnormal
protein.

Base pair substitutions
◦ Silent mutations – no
effect on encoded
protein due to
redundancy
◦ Missense – still codes
for an amino acid but
not the “right sense”.
◦ Nonsense* – a stop
codon in mid gene
* leads to nonfunctional
proteins

Base pair insertion mutations
◦ Addition or loss of base
pairs Disastrous
effect
◦ Frame-shift mutation – #
of nucleotides added or
deleted is not a multiple
of three severe
missense.
◦ why is a multiple of 3
okay?

Frame-shift mutation examples
◦ CAT ATE THE RAT becomes . . .
◦ CAR TAT ETH ERA T
◦ CAR TER ATE THE AT
Disease
related
mutation
statistics

Summary of transcription & translation