DNA acts as a template for protein production. It is transcribed into mRNA which is then translated into protein with the help of tRNA and ribosomes. Transcription occurs in the nucleus and copies DNA into RNA. Translation occurs in the cytoplasm and uses mRNA to assemble amino acids into a protein chain based on the genetic code. Gene expression can be regulated at the transcriptional or translational level to control the amount of protein produced.

1. Chapter 7
From DNA to Protein

2. DNA to Protein
• DNA acts as a “manager” in the process
of making proteins
• DNA is the template or starting
sequence that is copied into RNA that is
then used to make the protein

3. Central Dogma
• One gene – one protein

4. Central Dogma
• This is the same for bacteria to humans
• DNA is the genetic instruction or gene
• DNA  RNA is called Transcription
– RNA chain is called a transcript
• RNA  Protein is called Translation

5. Expression of
Genes
• Some genes are
transcribed in large
quantities because
we need large
amount of this
protein
• Some genes are
transcribed in small
quantities because
we need only a
small amount of
this protein

6. Transcription
• Copy the gene of interest into RNA which is
made up of nucleotides linked by
phosphodiester bonds – like DNA
• RNA differs from DNA
– Ribose is the sugar rather than deoxyribose –
ribonucleotides
– U instead of T; A, G and C the same
– Single stranded
• Can fold into a variety of shapes that allows RNA to
have structural and catalytic functions

7. RNA Differences

8. RNA Differences

9. Transcription
• Similarities to DNA replication
– Open and unwind a portion of the DNA
– 1 strand of the DNA acts as a template
– Complementary base-pairing with DNA
• Differences
– RNA strand does not stay paired with DNA
• DNA re-coils and RNA is single stranded
– RNA is shorter than DNA
• RNA is several 1000 bp or shorter whereas DNA is
250 million bp long

10. Template to Transcripts
• The RNA transcript is identical to the NON-
template strand with the exception of the T’s
becoming U’s

11. RNA Polymerase
• Catalyzes the formation of
the phosphodiester bonds
between the nucleotides
(sugar to phosphate)
• Uncoils the DNA, adds the
nucleotide one at a time
in the 5’ to 3’ fashion
• Uses the energy trapped
in the nucleotides
themselves to form the
new bonds

12. RNA Elongation
• Reads template 3’
to 5’
• Adds nucleotides 5’
to 3’ (5’ phosphate
to 3’ hydroxyl)
• Synthesis is the
same as the leading
strand of DNA

13. RNA Polymerase
• RNA is released so we can make many copies of
the gene, usually before the first one is done
– Can have multiple RNA polymerase molecules on a
gene at a time

14. Differences in
DNA and RNA Polymerases
• RNA polymerase adds ribonucleotides not
deoxynucleotides
• RNA polymerase does not have the ability to
proofread what they transcribe
• RNA polymerase can work without a primer
• RNA will have an error 1 in every 10,000
nucleotides (DNA is 1 in 10,000,000 nucleotides)

15. Types of RNA
• messenger RNA (mRNA) – codes for
proteins
• ribosomal RNA (rRNA) – forms the core of
the ribosomes, machinery for making
proteins
• transfer RNA (tRNA) – carries the amino
acid for the growing protein chain

16. DNA Transcription in Bacteria
• RNA polymerase must know where the start
of a gene is in order to copy it
• RNA polymerase has weak interactions with
the DNA unless it encounters a promoter
– A promoter is a specific sequence of nucleotides
that indicate the start site for RNA synthesis

17. RNA Synthesis
• RNA pol opens the
DNA double helix
and creates the
template
• RNA pol moves nt
by nt, unwinds the
DNA as it goes
• Will stop when it
encounters a STOP
codon, RNA pol
leaves, releasing the
RNA strand

18. Sigma () Factor
• Part of the bacterial RNA polymerase that
helps it recognize the promoter
• Released after about 10 nucleotides of RNA
are linked together
• Rejoins with a released RNA polymerase to
look for a new promoter

19. Start and Stop Sequences

20. DNA Transcribed
• The strand of DNA transcribed is dependent on which strand
the promoter is on
• Once RNA polymerase is bound to promoter, no option but to
transcribe the appropriate DNA strand
• Genes may be adjacent to one another or on opposite strands

21. Eukaryotic Transcription
• Transcription occurs in the nucleus in eukaryotes, nucleoid
in bacteria
• Translation occurs on ribosomes in the cytoplasm
• mRNA is transported out of nucleus through the nuclear
pores

22. RNA Processing
• Eukaryotic cells process the RNA in the
nucleus before it is moved to the cytoplasm
for protein synthesis
• The RNA that is the direct copy of the DNA is
the primary transcript
• 2 methods used to process primary transcripts
to increase the stability of mRNA being
exported to the cytoplasm
– RNA capping
– Polyadenylation

23. RNA Processing
• RNA capping happens at the 5’ end of the RNA, usually
adds a methylgaunosine shortly after RNA polymerase
makes the 5’ end of the primary transcript
• Polyadenylation modifies the 3’ end of the primary
transcript by the addition of a string of A’s

24. Coding and Non-coding Sequences
• In bacteria, the RNA made is translated to a protein
• In eukaryotic cells, the primary transcript is made of
coding sequences called exons and non-coding sequences
called introns
• It is the exons that make up the mRNA that gets translated
to a protein

25. RNA Splicing
• Responsible for the removal of the introns to create the
mRNA
• Introns contain sequences that act as cues for their removal
• Carried out by small nuclear riboprotein particles (snRNPs)

26. snRNPs
• snRNPs come together
and cut out the intron
and rejoin the ends of
the RNA
• Intron is removed as a
lariat – loop of RNA
like a cowboy rope

27. Benefits of Splicing
• Allows for genetic recombination
– Link exons from different genes together to create a new
mRNA
• Also allows for 1 primary transcript to encode for
multiple proteins by rearrangement of the exons

28. Summary

29. RNA to Protein
• Translation is the process of turning
mRNA into protein
• Translate from one “language” (mRNA
nucleotides) to a second “language”
(amino acids)
• Genetic code – nucleotide sequence
that is translated to amino acids of
the protein

30. Degenerate DNA Code
• Nucleotides read 3 at a time meaning that there
are 64 combinations for a codon (set of 3
nucleotides)
• Only 20 amino acids
– More than 1 codon per AA – degenerate code with the
exception of Met and Trp (least abundant AAs in
proteins)

31. Reading Frames
• Translation can occur in 1 of 3 possible reading
frames, dependent on where decoding starts in the
mRNA

32. Transfer RNA
Molecules
• Translation requires an
adaptor molecule that
recognizes the codon on
mRNA and at a distant site
carries the appropriate
amino acid
• Intra-strand base pairing
allows for this
characteristic shape
• Anticodon is opposite
from where the amino
acid is attached

33. Wobble Base Pairing
• Due to degenerate code for amino acids some tRNA
can recognize several codons because the 3rd spot
can wobble or be mismatched
• Allows for there only being 31 tRNA for the 61
codons

34. Attachment of AA to tRNA
• Aminoacyl-tRNA synthase is the enzyme
responsible for linking the amino acid to
the tRNA
• A specific enzyme for each amino acid and
not for the tRNA

35. 2 ‘Adaptors’ Translate
Genetic Code to Protein
1
 2

36. Ribosomes
• Complex machinery that
controls protein synthesis
• 2 subunits
– 1 large – catalyzes the peptide
bond formation
– 1 small – binds mRNA and tRNA
• Contains protein and RNA
– rRNA central to the catalytic
activity
• Folded structure is highly conserved
– Protein has less homology and
may not be as important

37. Ribosome Structures
• May be free in cytoplasm or attached to the ER
• Subunits made in the nucleus in the nucleolus and
transported to the cytoplasm

38. Ribosomal Subunits
• 1 large subunit – catalyzes the formation of the peptide bond
• 1 small subunit – matches the tRNA to the mRNA
• Moves along the mRNA adding amino acids to growing protein
chain

39. Ribosomal Movement
• 4 binding sites
– mRNA binding site
– Peptidyl-tRNA binding site (P-site)
• Holds tRNA attached to growing end of the peptide
– Aminoacyl-tRNA binding site (A-site)
• Holds the incoming AA
– Exit site (E-site)
E-site

40. 3 Step Elongation Phase
• Elongation is a cycle of events
• Step 1 – aminoacyl-tRNA comes into empty
A-site next to the occupied P-site; pairs with
the codon
• Step 2 – C’ end of peptide chain uncouples
from tRNA in P-site and links to AA in A-site
– Peptidyl transferase responsible for bond
formation
– Each AA added carries the energy for the addition
of the next AA
• Step 3 – peptidyl-tRNA moves to the P-site;
requires hydrolysis of GTP
– tRNA released back to the cytoplasmic pool

41. Initiation Process
• Determines whether mRNA is synthesized
and sets the reading frame that is used to
make the protein
• Initiation process brings the ribosomal
subunits together at the site where the
peptide should begin
• Initiator tRNA brings in Met
– Initiator tRNA is different than the tRNA that
adds other Met

42. Ribosomal Assembly
Initiation Phase
• Initiation factors (IFs) catalyze the steps –
not well defined
• Step 1 – small ribosomal subunit with the IF
finds the start codon –AUG
– Moves 5’ to 3’ on mRNA
– Initiator tRNA brings in the 1st AA which is
always Met and then can bind the mRNA
• Step 2 – IF leaves and then large subunit
can bind – protein synthesis continues
• Met is at the start of every protein until
post-translational modification takes place

43. Eukaryotic vs Procaryotic
• Procaryotic
– No CAP; have specific ribosome binding site upstream of AUG
– Polycistronic – multiple proteins from same mRNA
• Eucaryotic
– Monocistronic – one polypeptide per mRNA

44. Protein Release
• Protein released when a STOP codon
is encountered
– UAG, UAA, UGA (must know these
sequences!)
• Cytoplasmic release factors bind to
the stop codon that gets to the A-
site; alters the peptidyl transferase
and adds H2O instead of an AA
• Protein released and the ribosome
breaks into the 2 subunits to move
on to another mRNA

45. Polyribosomes
• As the ribosome
moves down the
mRNA, it allows for
the addition of
another ribosome and
the start of another
protein
• Each mRNA has
multiple ribosomes
attached,
polyribosome or
polysome

46. Regulation of Protein Synthesis
• Lifespan of proteins vary, need
method to remove old or damaged
proteins
• Enzymes that degrade proteins are
called proteases – process is called
proteolysis
• In the cytosol there are large
complexes of proteolytic enzymes
that remove damaged proteins
• Ubiquitin, small protein, is added as
a tag for disposal of protein

47. Protein Synthesis
• Protein synthesis takes the most energy
input of all the biosynthetic pathways
• 4 high-energy bonds required for each AA
addition
– 2 in charging the tRNA (adding AA)
– 2 in ribosomal activities (step 1 and step 3 of
elongation phase)

48. Summary

49. Ribozyme
• A RNA molecule can fold due
to its single stranded nature
and in folding can cause the
cleavage of other RNA
molecules
• A RNA molecule that functions
like an enzyme hence ribozyme
name