Transcript RNA Polymerase

Protein Synthesis
I. Protein Production
A. Background Info
1. DNA (in the nucleus) is the blueprint for
creating proteins
2. Ribosomes (in the cytoplasm) are where
ALL proteins are initially produced:
- proteins staying inside cell will be
completed here
- proteins to be exported or that become
lysosomes are transferred to ribosomes
on RER and synthesis is finished there
3. RNA carries out P.S. by acting as a
messenger b/n DNA & ribosomes.
B. RNA v. DNA
1. RNA is single-stranded
2. RNA is found in the nucleus & the cytoplasm
3. RNA is also a nucleic acid made up of
nucleotides consisting of 3 parts:
a. phosphate group
b. pentose sugar
c. nitrogenous base
HOWEVER…
Ribose
DEOXYribose
new base pairing
rule:
A=U
C=G
4. Three main kinds of RNA:
a. mRNA – carries info from DNA to ribosomes
b. tRNA – carries amino acid from cytoplasm to
ribosome
c. rRNA – help build ribosomes; binds mRNA
and tRNA together to make polypeptide chain
C. CENTRAL DOGMA
D. Protein Synthesis
1. Transcription - Takes place in the Nucleus
WHY?
 Taking info found in the DNA and turning
it into a molecule of mRNA
 As in replication, the DNA must unwind;
however, ONLY ONE strand of DNA
is used as a template, the other remains
untranscribed
*** DNA IS TRANSCRIBED 3’ to 5’
RNA IS SYNTHESIZED 5’ to 3’
a.Initiation
1. RNA Polymerase – the enzyme that binds
to DNA and transcribes it into mRNA.
2. Promoter - a specific sequence of nucleotides
on the DNA that tells RNA poly “bind here”
 this sequence is known as the
“TATA box” (~25 bases upstream from
the gene to be transcribed)
 RNA polymerase will orient itself here
 ~20 base pairs of DNA are unwound and
then RNA poly reaches start site and
begins transciption
b. Elongation
 RNA Polymerase moves along and “reads”
the DNA adding complimentary RNA
nucleotides (Chargoff’s base-pairing rules)
c. Termination (Fig. 1)
 RNA Poly continues until it reaches a “stop signal”
sequence
 RNA Poly will detach and release the new mRNA
 This new mRNA (i.e. transcript) now carries the
instructions for making proteins
mRNA leaves the nucleus and goes where?
To a ribosome in the cytoplasm
IMPORTANT: Before the new mRNA moves to
the cytoplasm, SPLICING occurs
1. INTRONS: Non-coding regions of DNA
 segments of DNA that are cut out b/c they
do not code for any part of a protein
2. EXONS: Coding regions of DNA
 segments of DNA that are EXpressed
 code for proteins
mRNA Processing (i.e splicing)
http://highered.mcgrawhill.com/sites/0072437316/student_view0/chapter15/animations.html
Promoter region
2. Translation
 translating the info on mRNA into amino
acids (i.e. polypeptide chain  protein)
 mRNA is translated 5’ to 3’
 based on the Genetic Code…
CODON - a series of 3 mRNA nucleotides that specify:
1. a particular amino acid
2. a “start signal” (only one)
3. a “stop signal” (3 different)
(64 possible codons)
Example:
If a gene is 99 DNA base pairs long ~~~~~~~~~ (99)
The mRNA is also 99 base pairs long ------------------ (99)
The protein will have 33 amino acids @@@@@@ (33)
99/3 = 33
 Other things needed for translation:
1. Ribosomes: contain two subunits,
large (heavy) and small (light)
Interesting fact:
- Ribosomes have 3 sections you APE:
A = Acceptor site: where tRNA enters
P = Peptidyl site: where one amino acid is
bonded to another in the polypeptide
chain
E = Exit site: where tRNA molecule leaves
after dropping off its amino acid
2. tRNA’s: carry the amino acids
- they contain a region called the ANTICODON:
a sequence of 3 nucleotides that is
complimentary to the codon on the mRNA
- where tRNA binds to mRNA
a.Initiation
 Ribosome subunits recognize and bind to
a recognition sequence on mRNA
 Ribosome then begins moving along mRNA
in a 5’ to 3’ direction
- translation initiates when ribosome reaches
the “start” codon (AUG)
 the first amino acid (METHIONINE) enters
the P site, the ONLY amino acid to do that.
b. Elongation
 The ribosome moves along the mRNA and
new amino acids (carried by tRNA) are
added forming a polypeptide chain
 Amino acids are linked by peptide bonds
c. Termination (Fig. 2)
 The ribosome reaches one of three “stop”
codons (i.e. there is no complimentary
tRNA anticodon)
 No more amino acids can be added so the
ribosome detaches & releases new protein
Signal sequence will determine what proteins are
finished being synthesized on the ribosomes of
the RER
http://highered.mcgrawhill.com/sites/0072437316/student_view0/chapter15/animations.html
II. Mutations – Changes in the nucleotide
sequence of DNA. Two general categories:
A. Point Mutations: mutations of single genes
1. base-substitutions (one base for another)
 two kinds:
a. transition:
purine for purine (AG)
pyrimidine for pyrimidine (CT)
b. transversion:
pyrimidine for purine (CA)
or vice versa
2. Frameshift Mutations
 involve the insertion or deletion of one or
more nucleotides from DNA
 causes a shift in the reading frame
 almost always lead to non-fxning protein
e.g. THE CAT ATE THE RAT
deletion of C
THE ATA TET HER AT
 base-substitutions & frameshifts are either:
1. silent – code for the same amino acid
2. missense – code for a different amino acid
3. nonsense – code for a stop codon
(can lead to nonfxning protein)
 Silent mutation
 Missense mutation
 Nonsense mutation
B. Chromosomal Mutations
 chromosomes may break during replication
and rejoin in abnormal ways
 4 specific types (examine during genetics)
Remember:
Changes to the amino acid sequence probably changes
the three-dimensional shape of the protein. Since
protein function if highly dependent on shape, this will
lead to impaired fxn or possibly complete nonfxn of the
protein!
If an individual inherits mutated genes for a single
protein, and if that protein is essential for life, the
individual may have seriously impaired health or may
even die
Examples: hemophilia, sickle-cell anemia, cystic fibrosis,
Huntington’s
III. Gene Expression
A. Gene Regulation, WHY?
--Why don’t organisms just express every gene in
their genome all the time?
--Bacteria (E. coli), for example, live in a wide
range of env’tal conditions and it is more
efficient to express only those genes that are
necessary for survival
--Remember, a high amount of NRG is involved
in gene expression (i.e. TXN & TLN)
--THUS, genes need to be regulated
B. How are genes regulated?
(Prokaryotic Mechanisms)
1. Genes responsible for a given cellular fxn
are organized into operons
2. These operons may be turned on (inducible)
or turned off (repressible) depending on the
situation
3. EXAMPLE: Lactose Metabolism (Inducible)
Tryptophan sythesis (repressible)
The Lac Operon: An Inducible System
Operator – the On/Off switch of a particular gene
Promoter – where RNA Polymerase binds to DNA to
begin transcription (1 per set of genes)
Repressor – Protein that blocks RNA Polymerase from
binding, thus NO txn & NO gene expression
(comes from repressor gene)
Structural Genes - Indicate the primary structure of a
proteins (i.e specific a.a. sequence)
DNA 
Situation #1 – Lactose IS present
 When Lactose is digested, it is broken down
as follows:
beta-galactosidase
Lactose ---------------------- glucose & galactose (major)
 Lactose is known as an inducer – a compound
that evokes synthesis of an enzyme
- in this example, lactose will induce the
production of enzymes Z, Y, and A
1. lactose (inducer) will bind to repressor &
cause a shape change
2. Repressor can no longer recognize the
operator binding site; switch is turned ON
3. RNA Polymerse can
bind to the operon’s
promoter
4. RNA Polymerase
begins to transcribe
the genes & genes
are then translated
5. The genes produce
enzymes (Z, Y, A)
that help break down
lactose
Situation #2: Lactose is NOT present
1. In the absence of lactose, there is no lactose to
bind to the Repressor enzyme & block it from
binding to operator
2. Thus, Repressor does bind to the operator and
the switch is left OFF
3. RNA Polymerase is now blocked from binding
to the lacPromoter
4. Thus NO TRANSCRIPTION of the genes
http://bcs.whfreeman.com/thelifewire/conte
nt/chp13/1302001.html
The Trp Operon: A Repressible System
 Sometimes tryptophan is present in high
concentrations (uh, THANKSGIVING, YUM!!!!!!)
so it is advantageous to stop making enzymes
for tryptophan synthesis
 These enzymes are said to be repressible
Situation #1: Tryptophan is NOT present
1. Repressor gene produces an inactive repressor
which cannot bind to operator (so stays ON)
2. RNA polymerase can bind to operator,
transcribe genes  enzymes will make Trp.
Situation #2: Tryptophan IS present
1. Tryptophan (i.e. co-repressor) will bind to
repressor protein and activates the repressor
2. Repressor binds to operator
3. RNA polymerase can’t bind to operator, thus
genes are not transcribed, no Trp made
http://bcs.whfreeman.com/thelifewir
e/content/chp13/1302002.html
SO, What is the difference b/n Inducible & Repressible
Systems? Summarize.
In inducible systems, a substance in the env’t
(i.e. the inducer) interacts with the repressor
making it incapable of binding to operator and
blocking transcription. (enzyme will be produced)
In Repressible systems, a substance in env’t
(i.e the corepressor) binds to repressor to make
it capable of binding to operator and blocking
transcription
C. Eukaryotic Gene Regulation/Expression
--Eukaryotes do not have a universal
mechanism (i.e operons) that controls the
activity of coding genes
--Rather, regulation is possible at any point in
the pathway b/n gene to functional protein