1. Protein Synthesis: Transcription and Translation

2. From nucleus to cytoplasm
transcription
DNA
mRNA
protein
translation
trait
nucleus
cytoplasm

3. Review
Central Dogma of Molecular Biology

4. DNA vs RNA DNA RNA Sugar: deoxyribose ribose Bonds with Adenine:
thymine
uracil
# of Strands:
two
one

5. Kinds of RNA
Messenger RNA (mRNA):copies DNA’s code & carries the genetic information to the ribosomes
Ribosomal RNA (rRNA):along with protein, makes up the ribosomes
Transfer RNA (tRNA): transfers amino acids to the ribosomes where proteins are synthesized

6. Transcription Making mRNA from DNA
ONE DNA strand is the template (pattern)
match bases
A: U
G : C
Enzyme
RNA polymerase

7. Sense vs. Antisense Strand
The antisense strand is the template strand during transcription, which is copied for translation into a protein.
a sense strand, or coding strand, is the strand of DNA running from 5' to 3' that is complementary to the antisense strand of DNA, which runs from 3' to 5'.
The sense strand is the strand of DNA that has the same sequence as the mRNA.

8. DNA contains codes

9. Transcription in prokaryotes
RNA polymerase recognizes a specific base sequence in the DNA called a promoter and binds to it.
The promoter identifies the start of a gene, which strand is to be copied, and the direction that it is to be copied.
RNA POLYMERASE- adds RNA nucleotides complimentary to the DNA template strand

10. Complimentary bases are assembled (U instead of T).
A termination code in the DNA indicates where transcription will stop in prokaryotes.
The mRNA produced is called a mRNA transcript.

11. Transcription Eukaryotes
Step 1: Transcription factors proteins bind and double stranded DNA unzips as RNA polymerase binds to promoter (TATAA sequence “TATA box” or other promoter) T

12. Matching bases of DNA & RNA
Step 2: Match RNA bases to DNA bases on one of the DNA strands. Built 5’ 3’, using DNA 3’-5’ template C U G A G U G U C U G C A A C U A A G C
RNA
polymerase U A G A C C T G G T A C A G C T A G T C A T C G T A C C G T

13. Matching bases of DNA & RNA
U instead of T is matched to A and the mRNA transcript is built 5’-3’
TACGCACA TTTA CGTA CG
DNA
AUGCGUGUAAAUGCAUGC
mRNA

14. Ending Transcription
Step 3: Transcription stops upon reading polyadenylation signal sequence.

15. RNA Polymerase
5’ 3’

16. mRNA Processing in Eukaryotes
After the DNA is transcribed into RNA, editing must be done to the nucleotide chain to make the RNA functional
Introns, non-functional segments of DNA are snipped out of the chain by snRNP’s (spliceosomes or ribonucleases)

17. mRNA Editing
Exons, segments of DNA that code for proteins, are then rejoined by the enzyme ligase
A guanine triphosphate cap is added to the 5” end of the newly copied mRNA
A poly A tail is added to the 3’ end of the RNA
The newly processed mRNA can then leave the nucleus through nuclear pores

18. Result of Transcription
New Transcript
Tail
CAP

19. mRNA Transcript
mRNA leaves the nucleus through its pores and goes to the ribosomes

20. How does mRNA code for proteins
mRNA leaves nucleus
mRNA goes to ribosomes in cytoplasm
Proteins built from instructions on mRNA
mRNA U C A G
How? aa

21. Translation

22. mRNA to protein = Translation
The working instructions  mRNA
The reader  ribosome
The transporter  transfer RNA (tRNA)
ribosome
mRNA U C A G aa
tRNA G U aa
tRNA U A C aa
tRNA G A C
tRNA aa A G U

23. Translation
Translation is the process of decoding the mRNA into a polypeptide chain
Ribosomes read mRNA three bases or 1 codon at a time and construct the proteins
The process begins by the mRNA leaving the nucleus through nuclear pores

24. Messenger RNA (mRNA) Carries the information for a specific protein
Made up of 500 to 1000 nucleotides long
Sequence of 3 bases called codon
AUG – methionine or start codon
UAA, UAG, or UGA – stop codons

25. Initiation and Termination Codes
An initiation code “start codon” signals the start of a genetic message. As the ribosome moves along a mRNA transcript, it will not begin synthesizing protein until it reaches an initiation code. (Ex. AUG)
Termination codes “stop codon” signal the end of the genetic message. Synthesis stops when the ribosome reaches a terminator codon.(Ex. UAA, UAG, UGA)

26. mRNA codes for proteins in triplets
TACGCACATTTACGTACGCGG
DNA
codon
ribosome
AUGCGUGUAAAUGCAUGCGCC
mRNA
AUGCGUGUAAAUGCAUGC ?
Met Arg Val Asn Ala Cys Ala
protein
Codon = block of 3 mRNA bases

27. The Genetic Code
Use the code by reading from the center to the outside
Example: AUG codes for Methionine
Genetic code contains a wobble at the third b.p.-there is redundancy in the genetic code, resulting in fewer errors.

28. Genetic Code- Another way

29. Name the Amino Acids
GGG?
UCA?
CAU?
GCA?
AAA?

30. Translation overview

31. Ribosomes Made of a large and small subunit
Composed of rRNA (40%) and proteins (60%)
Have three sites for tRNA attachment --- A –accepting site and P site- peptide bond
The tRNA exits from the E site

32. Transfer RNA (tRNA) Clover-leaf shape3’
Clover-leaf shape
Single stranded molecule with attachment site at 3’ end for an amino acid
Opposite end has three nucleotide bases called the anticodon 5’

33. Transfer RNA
The amino acid is bound to the 3’ end by a covalent ester bond through a condensation reaction
This is done by aminoacyl-tRNA synthetases or tRNA activating enzyme- one for every a.a.
tRNA is held together by intramolecular base pairing
There is a specific tRNA molecule for every codon or for every amino acid

34. Codons and Anticodons Recall a codon is made from 3 bases within mRNA
The 3 bases of an anticodon are complimentary to the 3 bases of a codon
Example: Codon ACU
Anticodon UGA
UGA
ACU

35. The Process

36. Step 1- Initiation
mRNA transcript start codon AUG attaches to the small ribosomal subunit
Small subunit attaches to large ribosomal subunit
mRNA transcript

37. tRNA Binding Steps
The amino acid will react with ATP and become activated (with aid of enzyme ATPase). The ATP lose energy in this process
2. The activated amino acid will then bind to the acceptor stem (3’) of its own tRNA by a covalent bond aided by tRNA synthetase
The bond between the tRNA and the a.a is broken when the peptide bond is formed

38. Ribosome “walks” the strand of mRNA 3 bases at a time 5’-3’
Step 2 - Elongation
As the ribosome moves, one tRNA carrying the Met amino acid moves into the A site, where the anticodon matches the mRNA transcript (AUG).
It then moves to the P site of the ribosome and another tRNA brings the next amino acid
Peptide bonds join the amino acids and leaves the P site tRNA empty (done by peptidyl transferase)
The empty tRNA is moved to the E site where it exits
Ribosome “walks” the strand of mRNA 3 bases at a time 5’-3’

39. 1. Transfer of proper tRNA to A-site of ribosome.
2. peptidyl transfer/
formation of peptide bond
3. translocation

40. Step 3-Termination When the ribosome reads a stop codon.
When this code (UGA, UAA, UAG) is reached protein translation is terminated.
Release factor binds
Polypeptide dissociates from ribosome
Occurs once per protein

41. End Product –The Protein!
The end products of protein synthesis is a primary structure of a protein
A sequence of amino acid bonded together by peptide bonds
aa1
aa2
aa3
aa4
aa5
aa200
aa199

42. Ribosomes E Site P Site A Site A U G C Large subunit mRNA
Small subunit

43. Initiation G aa2 A U U A C aa1 A U G C codon hydrogen bonds 2-tRNA
anticodon A U G C
hydrogen
bonds
codon
mRNA

44. Elongation G A aa3 peptide bond aa2 aa1 U A C G A U A U G C U A C U U
3-tRNA G A
aa3
peptide bond
aa2
aa1
1-tRNA
2-tRNA
anticodon U A C G A U A U G C U A C U U C G A
hydrogen
bonds
codon
mRNA

45. Ribosomes move over one codon
aa1
peptide bond
3-tRNA G A
aa3
aa2
1-tRNA U A C
(leaves)
2-tRNA G A U A U G C U A C U U C G A
mRNA
Ribosomes move over one codon

46. peptide bonds G C U aa4 aa1 aa2 aa3 G A U G A A A U G C U A C U U C G
4-tRNA G C U
aa4
aa1
aa2
aa3
2-tRNA
3-tRNA G A U G A A A U G C U A C U U C G A A C U
mRNA

47. Ribosomes move over one codon
peptide bonds
4-tRNA G C U
aa4
aa1
aa2
aa3
2-tRNA G A U
(leaves)
3-tRNA G A A A U G C U A C U U C G A A C U
mRNA
Ribosomes move over one codon

48. peptide bonds U G A aa5 aa1 aa2 aa4 aa3 G A A G C U G C U A C U U C G
5-tRNA
aa5
aa1
aa2
aa4
aa3
3-tRNA
4-tRNA G A A G C U G C U A C U U C G A A C U
mRNA

49. Ribosomes move over one codon
peptide bonds U G A
5-tRNA
aa5
aa1
aa2
aa3
aa4
3-tRNA G A A
4-tRNA G C U G C U A C U U C G A A C U
mRNA
Ribosomes move over one codon

50. Termination aa5 aa4 aa3 primary structure of a protein aa2 aa1 A C U C
terminator
or stop
codon
200-tRNA A C U C A U G U U U A G
mRNA

51. Mutation Frameshift (deletion)
A mutation is a change in amino acid sequence and can be either a;
Frameshift (deletion)
Original DNA: Frameshift mutation: 
THE BIG RED ANT ATE ONE FAT BUG THB IGR EDA NTA TEO NEF ATB UG?
Point Mutation
Original DNA: Point mutation:
THE BIG RED ANT ATE ONE FAT BUG THA BIG RED ANT ATE ONE FAT BUG

52. Silent, Missense, and Nonsense Mutations
 Three kinds of point mutations can occur. A mutation that results in an amino acid substitution is called a missense mutation. 
A mutation that results in a stop codon so that incomplete proteins are produced, it is called a nonsense mutation. 
A mutation that produces a functioning protein is called a silent mutation.

53. Terms Splicesomes- removes introns and ligase joins RNA polymerase-
adds RNA nucleotides complimentary to the DNA template strand
Transcription factors-
proteins that recognize specific sequences in DNA when making mRNA and help RNA polymerase bind
ATPase-
converts ATP to ADP and releases energy to do work ( used to bond tRNA to mRNA and GTPase also used when adding a.a to tRNA)
tRNA synthetase (tRNA activating enzyme) -
enzyme that catalyzes the esterification of a specific amino acid to a specific tRNA
Peptidyltransferase-
forms peptide bonds and helps with protein release
recognizes tRNA code and adds specific a.a. (a covalent bond)
Protein release factors-
Helps release peptide during termination of translation by recognizing the termination codon or stop codon in an mRNA sequence.

54. Question:
What would be the complimentary RNA strand for the following DNA sequence?
DNA 5’-GCGTATG-3’

55. Answer:
DNA 5’-GCGTATG-3’
RNA 3’-CGCAUAC-5’

56. Exercise - Understanding DNA, mRNA, tRNA, and protein.
Template (anti-sense) strand
GGG
Non-template strand
TAC
mRNA
 CUU
CCU
tRNA
UCG
Amino Acid
Leu