1. Quiz
Nucleotide?
Added to which end?
Lagging strand?

2. DNA pol III synthesizes leading strand continuously3 5
Parental DNA
DNA pol III starts DNA synthesis at 3 end of primer, continues in 5  3 direction 5 3 5
Lagging strand synthesized in short Okazaki fragments, later joined by DNA ligase
Primase synthesizes a short RNA primer 3 5

3. Chapter 17
From Gene to Protein

4. DNA > RNA > “Protein”
Big Questions
How does your body produce insulin?
How does your offspring know how to make insulin?
Central “Dogma”
DNA > RNA > “Protein”
(Info > Message > Product)

5. Chapter 17 Study Guide Questions
The Connection between Genes and Proteins
1. Explain how RNA differs from DNA.
2. Briefly explain how information flows from gene to protein. Is the central dogma ever violated?
3. Distinguish between transcription and translation.
4. Compare where transcription and translation occur in bacteria and in eukaryotes.
5. Define “codon” and explain the relationship between the linear sequence of codons on mRNA and the linear sequence of amino acids in a polypeptide.
6. Explain what it means to say that the genetic code is redundant and unambiguous.
7. Explain the significance of the reading frame during translation.

6. Chapter 17 Study Guide Questions
The Synthesis and Processing of RNA
8. Explain how RNA polymerase recognizes where transcription should begin. Describe the role of the promoter, the terminator, and the transcription unit.
9. Explain the general process of transcription, including the three major steps of initiation, elongation, and termination.
10. Explain how RNA is modified after transcription in eukaryotic cells.
11. Define and explain the role of ribozymes. What three properties allow some RNA molecules to function as ribozymes?
12. Describe the functional and evolutionary significance of introns.
13. Explain why, due to alternative RNA splicing, the number of different protein products an organism can produce is much greater than its number of genes.

7. Chapter 17 Study Guide Questions
The Synthesis of Protein 14. Describe the structure and function of tRNA. 15. Explain how tRNA is joined to the appropriate amino acid. 16. Describe the structure and functions of ribosomes. 17. Describe the process of translation (including initiation, elongation, and termination) and explain which enzymes, protein factors, and energy sources are needed for each stage. 18. Describe the significance of polyribosomes. 19. Explain what determines the primary structure of a protein and describe how a polypeptide must be modified before it becomes fully functional. 20. Define “point mutations”. Distinguish between base-pair substitutions and base-pair insertions. Give an example of each and note the significance of such changes.

8. Today’s Tip: Ricin will kill you
Georgi Ivanov Markov
Inhibits ribosome translation of mRNA

9. 1. Explain how RNA differs from DNA.
Single-stranded
AUCG
Ribose
DNA
Double-stranded
ATCG
Deoxyribose

10. 2. Briefly explain how information flows from gene to protein
2. Briefly explain how information flows from gene to protein. Is the central dogma ever violated?
Genes
Specific sequences of nucleotides
Proteins
Instructions carried in genes
Gene expression
DNA directs protein synthesis
Includes two stages:
transcription and translation
Cellular chain of command:
DNA RNA protein

11. 3. Distinguish between transcription and translation.
synthesis of RNA under the direction of DNA
Messenger RNA (mRNA)
Product of transcription
Translation
synthesis of a polypeptide
occurs under the direction of mRNA
_________are the sites of translation

12. 4. Compare where transcription and translation occur in bacteria and in eukaryotes.
Prokaryotes
Transcription – cytoplasm
Translation – cytoplasm
Eukaryotes
Transcription – Nucleus

13. TRANSCRIPTION Prokaryotic cellLE
DNA
TRANSCRIPTION
Prokaryotic cell

14. DNA TRANSCRIPTION mRNA Ribosome Prokaryotic cell PolypeptideLE
DNA
TRANSCRIPTION
mRNA
Ribosome
Prokaryotic cell
Polypeptide
Prokaryotic cell

15. LE 17-3-3 DNA TRANSCRIPTION mRNA Ribosome TRANSLATION Polypeptide
Prokaryotic cell
Nuclear
envelope
TRANSCRIPTION
DNA
Eukaryotic cell

16. LE 17-3-4 DNA TRANSCRIPTION mRNA Ribosome TRANSLATION Polypeptide
Prokaryotic cell
Nuclear
envelope
TRANSCRIPTION
DNA
Pre-mRNA
RNA PROCESSING
mRNA
Eukaryotic cell

17. LE 17-3-5 DNA TRANSCRIPTION mRNA Ribosome TRANSLATION Polypeptide
Prokaryotic cell
Nuclear
envelope
TRANSCRIPTION
DNA
Pre-mRNA
RNA PROCESSING
mRNA
Ribosome
TRANSLATION
Polypeptide
Eukaryotic cell

18. 5. Define “codon” and explain the relationship between the linear sequence of codons on mRNA and the linear sequence of amino acids in a polypeptide.
Codons
mRNA base triplets
specifies specific amino acid
amino acid placed at the corresponding position along a polypeptide
Template strand
provides a template for ordering the sequence of nucleotides in an RNA transcript

19. 5. Define “codon” and explain the relationship between the linear sequence of codons on mRNA and the linear sequence of amino acids in a polypeptide.
Triplet code
nonoverlapping
three-nucleotide words
code for amino acids
Example:
AGT on DNA strand
results in the placement of the amino acid serine
at the corresponding position of the polypeptide to be produced

20. 6. Explain what it means to say that the genetic code is redundant and unambiguous.
How many amino acids? 20
How many codons? 64
What does that imply?

21. 6. Explain what it means to say that the genetic code is redundant and unambiguous.
Redundant but not ambiguous (?)
1. One amino acid can have more than one codon
e.g., UCU, UCC, UCA, etc., all code for Serine
2. No codon specifies more than one amino acid

22. 7. Explain the significance of the reading frame during translation.
The fat cat ate the rat
Reading Frame
Codons must be read in the correct reading frame
What happens if you are off by one letter?
Frame Shift Error

23. Practice What amino acid does CAC code for? AGA? CGU?
Three code for stop
AUG codes for begin

24. Evolution of the Genetic Code
The genetic code is nearly universal
What does that mean?
shared by the simplest bacteria to the most complex animals
What does that imply?
Luciferase

25. 9. Explain the general process of transcription, including the three major steps of initiation, elongation, and termination.
The three stages of transcription:
1) Initiation
2) Elongation
3) Termination

26. Molecular Components of Transcription
RNA Polymerase
pries the DNA strands apart and hooks together the RNA nucleotides
RNA synthesis follows the same base-pairing rules as DNA
except __?__ substitutes for thymine

27. Animation: Transcription
8. Explain how RNA polymerase recognizes where transcription should begin. Describe the role of the promoter, the terminator, and the transcription unit.
Initiation
Promoter
DNA sequence where RNA polymerase attaches
Transcription Unit
stretch of DNA that is transcribed
Terminator
DNA sequence found only on prokaryotes
Animation: Transcription

28. Promoter Transcription unit 5¢ 3¢ 3¢ 5 DNA Start point RNA polymerase
LE 17-7
Promoter
Transcription unit 5¢ 3¢ 3¢ 5
DNA
Start point
RNA polymerase
Initiation 5¢ 3¢ 3¢ 5
RNA
tran-
script
Template strand
of DNA
Unwound
DNA
Elongation
Rewound
DNA 5¢ 3¢ 3¢ 3¢ 5 5¢
RNA
transcript

29. LE 17-7 Promoter Transcription unit 5 3 3¢ 5¢ DNA Start point
RNA polymerase
Initiation 5¢ 3¢ 3¢ 5¢
RNA
tran-
script
Template strand
of DNA
Unwound
DNA
Elongation
Rewound
DNA 5¢ 3¢ 3¢ 3¢ 5¢ 5¢
RNA
transcript
Termination 5¢ 3¢ 3¢ 5¢ 5¢ 3¢
Completed RNA transcript

30. Elongation of the RNA Strand
RNA polymerase
untwists the double helix
10 to 20 bases at a time
40 nucleotides per second (eukaryotes)
can be transcribed simultaneously by several RNA polymerases
Why?

31. Termination of Transcription
Mechanisms of termination are different in prokaryotes and eukaryotes
Prokaryotes
polymerase stops transcription at the end of the terminator sequence
Eukaryotes
polymerase continues transcription after the pre- mRNA is cleaved
Polymerase eventually falls off the DNA

32. 10. Explain how RNA is modified after transcription in eukaryotic cells.
Does the pre-mRNA now leave the nucleus?
How is the pre-mRNA modified?

33. Alteration of mRNA Ends
Each end of a pre-mRNA molecule is modified in a particular way:
The 5 end receives a modified nucleotide cap
The 3 end gets a poly-A tail
These modifications share several functions:
They seem to facilitate the export of mRNA
They protect mRNA from hydrolytic enzymes
They help ribosomes attach to the 5’ end

34. Split Genes and RNA Splicing
Intervening sequences (Introns)
long noncoding stretches - removed
Exons
shorter coding sections – will be expressed
RNA splicing
removes introns and joins exons
creating an mRNA molecule with a continuous coding sequence

35. RNA duplicating RNA, a step closer to the origin of life By Yun Xie |
NASA JPL
According to the “RNA world” model of life's origin, RNA performed all of the operations that are essential to life. RNA alone passed on genetic information and catalyzed the reactions of basic metabolism; DNA and proteins were not in the picture. The RNA world hypothesis is an appealingly simple model for simple early life forms, since it allows the complex array of biochemical interactions among proteins, DNA, and RNA to evolve gradually.
Our current natural world no longer uses RNA enzymes that act on their own to perform most biological functions. To better understand ancient RNA enzymes, modern scientists have to rely on proxies, like engineered RNA "ribozymes" that have catalytic functions without the need for proteins. However, scientists have had trouble creating a proxy for the first self-replicating molecule, or even an RNA ribozyme that can copy an RNA that's long enough to have further biological functions. Aniela Wochner and her coauthors have overcome that difficulty. In a recent issue of Science, they report the creation of an RNA ribozyme that synthesizes complex RNAs, including RNAs that act as ribozymes and perform a biological function.
Previously, the leading RNA polymerase ribozyme, called R18, could only transcribe RNAs up to 14 bases long (as a frame of reference, R18 itself is about 196 bases long). It was also highly template-dependent, meaning it could only copy certain sequences of RNA. To establish early life on Earth, a ribozyme would need to be able to make a variety of RNA sequences of adequate length, including something long enough to synthesize itself. Wochner and her colleagues sought to engineer a superior RNA ribozyme by modifying R18.
First, they wanted to improve the interactions among the template RNA, the ribozyme, and the primer sequence that starts the copying. To make RNA, the ribozyme has to recognize the primer and the template, which base-pair with one another. Then, the ribozyme catalyzes the addition of new bases onto the primer, making an RNA sequence that is complementary to the template. 
Scientists have proposed that the one end of the R18 ribozyme interacts with the primer and template. So, Wochner and her colleagues appended a random sequence into the 5’-end of R18 and selected for improved RNA polymerase activity.
They found one ribozyme (named C19) that did better than R18 on a specific, short template, but it didn't work well on longer templates. They further modified C19 by making truncated variants of its sequence and screened for improved activity on longer templates. They found one variant (the ribozyme tC19) that can extend primers by up to 95 bases with favorable templates.
The final obstacle was the fact that it only worked well on favorable template sequences—the researchers wanted a ribozyme that will be able to copy a diverse range of RNA templates, not just a few favorable ones. To find one, they made 50 million randomly mutated R18 sequences, did numerous rounds of selections, and found a combination of mutations that improved the recognition of diverse templates. They applied those mutations to the tC19 ribozyme, creating the RNA polymerase ribozyme tC19Z (198-bases long).
Ribozyme tC19Z synthesizes longer RNA sequences and can work with a greater range of primer-template combinations than any of the previous ribozymes. Wochner and her colleagues were able to use tC19Z to synthesize a minimal version of the hammerhead ribozyme (an RNA that binds to and cleaves an RNA substrate). The synthesized hammerhead ribozyme had catalytic activity, as it was able to cleave an RNA substrate at the expected location in the sequence.
Wochner and her coauthors have significantly expanded our abilities to engineer RNA polymerase ribozymes; however, further improvements are still necessary. For example, tC19Z probably cannot synthesize something of its own size in a reasonable amount of time. Nevertheless, it's impressive that the researchers were able to select for such drastic improvements on R18, as the sequence hasn't seen a significant upgrade since its creation almost a decade ago. Their work lead us closer to understanding ribozymes that could have existed in early Earth.
Science, DOI: /science (About DOIs)

36. Concept 17.4: Translation is the RNA-directed synthesis of a polypeptide: a closer look
Transfer RNA (tRNA)
translates an mRNA message into protein
Molecules of tRNA are not identical:
Each carries a specific amino acid on one end
Each has an anticodon on the other end

37. 14. Describe the structure and function of tRNA.
tRNA molecule
single RNA strand
~ 80 nucleotides long
cloverleaf shape
twists and folds into a three-dimensional molecule
How?
hydrogen bonds A C C

38. “problems”
A. How do you make sure that the correct amino acid to connects to the correct tRNA?
B. How do you make sure that the correct tRNA matches up with the correct codon?

39. 15. Explain how tRNA is joined to the appropriate amino acid.
Accurate translation requires two steps:
First step:
correct match between a tRNA and an amino acid
done by the enzyme aminoacyl-tRNA synthetase (20 types)
Second step:
- correct match between the tRNA anticodon and an mRNA codon

40. 16. Describe the structure and functions of ribosomes.
facilitate specific coupling of tRNA anticodons with mRNA codons in protein synthesis
Two ribosomal subunits (large and small)
made of proteins and ribosomal RNA (rRNA)

41. You have brucellosis. What do you do?
Tetracycline:
works by binding specifically to the 30S ribosome of the bacteria
preventing attachment of the aminoacyl tRNA to the RNA-ribosome complex

42. A ribosome has three binding sites for tRNA:
The P site
holds the growing polypeptide chain
The A site
holds the next amino acid to be added
The E site
is the exit site, where discharged tRNAs leave the ribosome

43. The three stages of translation: Initiation Elongation Termination
17. Describe the process of translation (including initiation, elongation, and termination) and explain which enzymes, protein factors, and energy sources are needed for each stage.
The three stages of translation:
Initiation
Elongation
Termination

44. Ribosome Association and Initiation of Translation
Initiation stage
brings together mRNA, a tRNA with the first amino acid, and the two ribosomal subunits

45. Ribosome Association and Initiation of Translation
First, a small ribosomal subunit binds with mRNA and a special initiator tRNA

46. Ribosome Association and Initiation of Translation
Then the small subunit moves along the mRNA until it reaches the start codon (AUG)

47. Ribosome Association and Initiation of Translation
Proteins called initiation factors bring in the large subunit so the initiator tRNA occupies the P site

48. Elongation of the Polypeptide Chain
Amino acids added one by one
Each addition involves proteins called elongation factors
Occurs in three steps:
codon recognition
peptide bond formation
translocation

49. 2 -peptide bond formation 3- translocation
LE 17-18
Amino end
of polypeptide E 3¢
mRNA P
site A
site
Ribosome ready for
next aminoacyl tRNA 5¢ 2
GTP
2 GDP E E P A P A
GDP
GTP
three steps:
1- codon recognition
2 -peptide bond formation
3- translocation E P A

50. Termination of Translation
LE 17-19
Termination of Translation
Release
factor
Stop codon
(UAG, UAA, or UGA) 5¢ 3¢
Free
polypeptide
When a ribosome reaches a stop
codon on mRNA, the A site of the
ribosome accepts a protein called
a release factor instead of tRNA. 3¢
The release factor hydrolyzes the
bond between the tRNA in the
P site and the last amino acid of the
polypeptide chain. The polypeptide
is thus freed from the ribosome.
The two ribosomal subunits
and the other components
of the assembly dissociate.
Ribosome acts as one large Ribozyme

51. 18. Describe the significance of polyribosomes.
make many copies of a polypeptide very quickly

52. 19. Explain what determines the primary structure of a protein and describe how a polypeptide must be modified before it becomes fully functional.
Primary structure
Amino acid sequence
Polypeptide chains are modified after translation
Post-translational modifications
Removal of amino acids
AUG
Splitting of polypeptide chain
insulin

53. Targeting Polypeptides to Specific Locations
Two populations of ribosomes are evident in cells:
free ribsomes (in the cytosol)
bound ribosomes (attached to the ER)
Free ribosomes mostly synthesize proteins that function in the cytosol

54. Targeting Polypeptides to Specific Locations
Bound ribosomes:
endomembrane system
secreted from the cell
Ribosomes are identical and can switch from free to bound

55. Concept 17.5: RNA plays multiple roles in the cell: a review
Type of RNA
Functions
Messenger RNA (mRNA)
Carries information specifying amino acid sequences of proteins from DNA to ribosomes
Transfer RNA (tRNA)
Serves as adapter molecule in protein synthesis; translates mRNA codons into amino acids
Ribosomal RNA (rRNA)
Plays catalytic (ribozyme) roles and structural roles in ribosomes

56. Type of RNA
Functions
Primary transcript
Serves as a precursor to mRNA, rRNA, or tRNA, before being processed by splicing or cleavage

57. Concept 17.6: Comparing gene expression in prokaryotes and eukaryotes reveals key differences
Prokaryotic cells:
lack a nuclear envelope
allows translation to begin while transcription progresses
Eukaryotic cell:
The nuclear envelope separates transcription from translation
Extensive RNA processing occurs in the nucleus

58. Coupled transcription and translation in bacteria
RNA polymerase
DNA
mRNA
Polyribosome
Direction of
transcription
0.25 mm
RNA
polymerase
DNA
Polyribosome
Polypeptide
(amino end)
Ribosome
mRNA (5¢ end)
Coupled transcription and translation in bacteria

59. changes in the genetic material of a cell or virus
20. Define “point mutations”. Distinguish between base-pair substitutions and base-pair insertions. Give an example of each and note the significance of such changes.
Mutations
changes in the genetic material of a cell or virus
Point mutations
chemical changes in just one base pair of a gene
The change of a single nucleotide in a DNA template strand leads to production of an abnormal protein

60. Wild-type hemoglobin DNA Mutant hemoglobin DNA
LE 17-23
Wild-type hemoglobin DNA
Mutant hemoglobin DNA 3¢ 5¢ 3¢ 5¢
mRNA
mRNA 5¢ 3¢ 5¢ 3¢
Normal hemoglobin
Sickle-cell hemoglobin

61. Substitutions Base-pair substitution
replaces one nucleotide and its partner with another pair of nucleotides
can cause missense or nonsense mutations
Missense
mutations still code for an amino acid, but not necessarily the right amino acid
Nonsense mutations
change an amino acid codon into a stop codon, nearly always leading to a nonfunctional protein
Missense mutations are more common

62. Insertions and Deletions
additions or losses of nucleotide pairs in a gene
have a disastrous effect on the resulting protein more often than substitutions do
may alter the reading frame, producing a frameshift mutation

63. Mutagens Spontaneous mutations
may occur during DNA replication, recombination, or repair
Mutagens
physical or chemical agents that can cause mutations
Mutagenic radiation, ultraviolet light (physical)
Cancer-causing chemicals