1. Translation and Proteins
Active Lecture PowerPoint® Presentation for Essentials of Genetics Seventh Edition Klug, Cummings, Spencer, Palladino
Chapter 13
Translation and Proteins
Copyright © 2010 Pearson Education, Inc.

2. Outline Overview of translation Components of translation machinery
The process of translation
Eukaryotic translation
Relationship between genes and proteins
Protein structure
Summary (animation)

3. Gene Expression Translation Step 1 Transcription Step 2
Why is step 2 called translation?
This second phase of protein synthesis is called translation because language of RNA is converted to language of proteins. Translating from a language written in four base alphabet into a second language written in a alphabet of 20 amino acids. DNA and RNA are made up of nucleotide letters, where as proteins are made out of amino acid letters.
RNA carries the DNA’s protein building instructions from the nucleus to the cytoplasm. RNA is a true messenger in this sense.
Step 2
Translation

4. mRNA

5. Components of Translation Machinery
mRNA transcribed from DNA
Amino acids
tRNAs
Ribosomes
mRNA transcribed from DNA
Amino acids are the building blocks of proteins
tRNA or transfer RNA bring the amino acids, There is one tRNA for each one of the 20 amino acids. So tRNAs provide escort service to amino acids.
Finally, ribosomes are the sites of protein synthesis in cells. These are mobile protein synthesising factories.

6. Translation - Components
Codon
mRNA carrying the message (or instructions for the protein) from DNA.
Two ribosomal subunits coming together to initiate translation
Once the mRNA is docked at a ribosome, its bases are read three at a time. We call this a codon.
tRNA brings the amino acid to the ribosomes. Three bases complementary to the codon is located in tRNA. This is called the anticodon. Anticodon is located in one end of the tRNA molecule, the other end has the corresponding amino acid; Therefore, tRNA is the actual translator or some times it is called the adapter molecule.

7. Ribosome Location of protein synthesis in cells
Composed of RNA and proteins
Composed of a large and a small subunit
Prokaryotes 50S and 30S
Eukaryotes 60S and 40S
Ribosome is much larger and complex than the machinary required for DNA and RNA synthesis

8. FIGURE 13-1
A comparison of the components in prokaryotic and eukaryotic ribosomes.
Ribosomes consist of ribosomal proteins and ribosomal RNAs (rRNAs) and have a large subunit and a small subunit
The rRNAs provide for all the important catalytic functions associated with translation.
rRNA genes (rDNA) are moderately repetitive and tandemly repeated.

9. Chapter 13 Opener
Crystal structure of a Thermus thermophilus 70S ribosome containing three bound transfer RNAs. Reprinted from the front cover of Science, Vol. 292, May 4, 2001.

10. tRNA (Clover leaf model) FIGURE 13-3
Holley's two-dimensional cloverleaf model of transfer RNA. Blocks represent nitrogenous bases.
tRNAs serve as adaptor molecules to adapt the triplet codons in mRNA to the correct amino acid.
tRNAs are 75–90 nucleotides long and contain posttranscriptionally modified bases.
The two-dimensional structure of tRNAs is a cloverleaf
A tRNA has an anticodon that base-pairs with the codon in the mRNA.
The corresponding amino acid is bound to the CCA sequence at the 3' end of all tRNAs.

11. FIGURE 13-4
A three-dimensional model of transfer RNA.

12. Charging tRNA by Aminoacyl tRNA Synthetase
FIGURE 13-5
Steps involved in charging tRNA. The superscript x denotes that only the corresponding specific tRNA and specific aminoacyl tRNA synthetase enzyme are involved in the charging process for each amino acid.
Aminoacyl tRNA synthetase charges (activates) tRNAs with the appropriate amino acid. There are 20 different aminoacyl tRNA synthetase enzymes.

13. Translation of mRNA Can Be Divided into Three Steps
Initiation
Elongation
Termination

14. Various Proteins Assist Translation
TABLE 13.1
Various Protein Factors Involved During Translation in E. coli
These are called Initiation Factors, Elongation Factors and Release Factors.

15. Ribosome Binding Sites
The ribosome has three binding sites for tRNA.
A site: Acceptor site
tRNA bringing amino acid binds
P site: Peptidyl site
tRNA carrying the peptide chain binds
E site: Exit site
First amino acid containing tRNA binds straight to the P site.

16. FIGURE 13-6 part 1
Initiation of translation. The components are depicted at the left of the figure.

17. Translation Initiation
Small subunit of the ribosome binds to the start codon of mRNA. Initiation factors help.
The first amino acid attached to an initiator tRNA, binds to mRNA
The first amino acid
Prokaryotes: Formyl-methionine (f-Met)
Eukaryotes: methionine
The large subunit of the ribosome binds
First amino acid containing tRNA binds straight to the P site.

18. Translation Initiation
FIGURE 13-6 part 2
Initiation of translation. The components are depicted at the left of the figure.

19. Translation Elongation
Ribosome advances one codon at a time
tRNA in A site is translocated to the P site so that the A site is available for the next charged tRNA to bind
A peptide bond is formed between amino acid in A site and the amino acid in the P site by peptidyl transferase, a ribozyme composed of RNA in the large subunit

20. FIGURE 13-7 part 1
Elongation of the growing polypeptide chain during translation.
Elongation requires both ribosomal subunits assembled with the mRNA to form the P (peptidyl) site and A (aminoacyl) site.
The charged tRNAs enter the A site, and peptidyl transferase catalyzes peptide bond formation between the amino acid on the tRNA at the A site and the growing peptide chain bound to the tRNA in the P site.
The uncharged tRNA moves to the E (exit) site, and the tRNA bound to the peptide chain moves to the P site.

21. FIGURE 13-7 part 2
Elongation of the growing polypeptide chain during translation.

22. FIGURE 13-7 part 3
Elongation of the growing polypeptide chain during translation.

23. FIGURE 13-7 part 4
Elongation of the growing polypeptide chain during translation.

24. FIGURE 13-7 part 5
Elongation of the growing polypeptide chain during translation.

25. Translation Termination
Ribosome binds to the stop codon of mRNA.
Ribosome releases mRNA & the polypeptide chain synthesized
Release factors assist this
Ribosome subunits dissociate

26. FIGURE 13-7 part 6
Elongation of the growing polypeptide chain during translation.

27. FIGURE 13-8
Termination of the process of translation.
Termination is signaled by a stop codon (UAG, UAA, UGA) in the A site.
GTP-dependent release factors cleave the polypeptide chain from the tRNA and release it from the translation complex.

28. Translation is More Complex in Eukaryotes
the ribosomes are larger than in bacteria
transcription and translation are spatially and temporally separated
ribosomes scan for the initiator tRNA that is in the proper context, as identified by the Kozak sequence
translation generally requires more factors for initiation, elongation, and termination than translation in bacteria does
Some ribosomes are not free-floating as in bacteria but instead are associated with the endoplasmic reticulum.

29. Relationship between Genes & Proteins
George Beadle (1933) provided evidence that genes are directly responsible for enzymes.
Work by Beadle & Edward Tatum on Neurospora led to one gene:one enzyme hypothesis
Studies on human hemoglobin established one gene: one polypeptide hypothesis
Because not all proteins are enzymes and some proteins have more than one subunit, the one-gene:one-enzyme hypothesis was modified to one-gene:one-protein and then to one-gene:one-polypeptide chain.

30. Experiments of Beadle & Tatum D A B C
Biochemical Pathways D A B C
Enzyme 1
Enzyme 2
Enzyme 3
Defective enzyme is a result of mutation in the gene
Hypothesis: One Gene - One Enzyme

31. Sickle-cell Anemia FIGURE 13-13
A comparison of erythrocytes from (a) healthy individuals, and (b) those with sickle-cell anemia.
First direct evidence that genes specify proteins other than enzymes came from work on human disorder sickle cell anemia.
Hemoglobin binds to and transports oxygen, which is essential for cellular metabolism.

32. Sickle-cell Anemia
Patients suffer from lack of oxygen to tissue and severe damage
Inherited as a recessive Mendelian trait Affected individuals are homozygous for Hbs
Heterozygots (HbAHbS) are carriers but not largely affected
HbS hemogloblin has a different rate of electrophoretic migration than HbA hemoglobin
The kidney, muscle, joints, brain, GI tract, lungs can be affected. Can be fatal, if left untreated.
Sickle-cell anemia is a recessive genetic disease in which afflicted individuals are homozygous for the HbS hemoglobin allele. Heterozygotes are carriers of the affected gene but are largely unaffected.
Fingerprinting demonstrated that the HbS and HbA hemoglobins differ by a single peptide fragment

33. FIGURE 13-14
Investigation of hemoglobin derived from HbAHbA, HbAHbS and HbSHbS individuals by using electrophoresis, protein fingerprinting, and amino acid analysis. Hemoglobin from individuals with sickle-cell anemia (HbSHbS) (a) migrates differently in an electrophoretic field, (b) shows an altered peptide in fingerprint analysis, and (c) shows an altered amino acid, valine, at the sixth position in the chain. During electrophoresis, heterozygotes (HbAHbS) reveal both forms of hemoglobin.

34. Protein Structure (self study)
Four levels of protein structure
Primary structure (amino acid chain)
Secondary structure (α-helices & β-sheets)
Tertiary structure (a single subunit)
Quarternary structure (group of subunits)

35. FIGURE part 1
Chemical structures and designations of the 20 amino acids found in living organisms, divided into four major categories. Each amino acid has two abbreviations; that is, alanine is designated either ala or A (a universal nomenclature).

36. FIGURE 13-15
Chemical structures and designations of the 20 amino acids found in living organisms, divided into four major categories. Each amino acid has two abbreviations; that is, alanine is designated either ala or A (a universal nomenclature).
Following translation, polypeptides fold up and assume higher order structures, and they may interact with other polypeptides.
Amino acids all have a carboxyl group, an amino group, and an R (radical) group bound to a central carbon atom. The R group of an amino acid confers specific chemical properties

37. FIGURE 13-16
Peptide bond formation between two amino acids, resulting from a dehydration reaction.
A peptide bond forms by a dehydration reaction between the carboxyl group of one amino acid and the amino group of another.

38. FIGURE 13-17
(a) The righthanded alpha helix, which represents one form of secondary structure of a polypeptide chain. (b) The beta-pleated sheet, an alternative form of secondary structure of polypeptide chains. To maintain clarity, not all atoms are shown.

39. Tertiary Level Protein
Respiratory
pigment
myoglobin
FIGURE 13-18
The tertiary level of protein structure in a respiratory pigment, myoglobin. The bound oxygen atom is shown in red.

40. Quarternary Level Protein
Hemoglobin
FIGURE 13-19
The quaternary level of protein structure as seen in hemoglobin. Four chains (two α and two β) interact with four heme groups to form the functional molecule.

41. Diversity in Protein Function
Proteins play diverse roles in the body
Examples: Hemoglobin, Collagen, Keratin, Actin, Myosin, Immunoglobulins, Transport proteins, some hormones and their receptors, histones, enzymes
Diversity in protein function is directly related to the structure of the molecule
Hemoglobin binds to and transports oxygen, which is essential for cellular metabolism.
Collagen and keratin are structural proteins.
Actin and myosin are contractile proteins, found in abundance in muscle tissue.
Other examples are the immunoglobulins, which function in the immune system of vertebrates; transport proteins, involved in movement of molecules across membranes; some of the hormones and their receptors, which regulate various types of chemical activity; and histones, which bind to DNA in eukaryotic organisms.

42. Important features of proteins
Some proteins may be posttranslationally modified
Protein function is directly related to the structure
Proteins consist of functional domains
Protein molecules may have domains that fold independently of the rest of the protein into stable, unique conformations. Different protein domains impart different functional capabilities.

43. Genetics, Technology & Society p 297
Prions (protenaceous infectious agents)
Mad cow
Creutzfeldt –Jakob disease
Kuru disease
Scrapie in sheep
BSE in cattle
Prions consist of protein only.
Prion protein and the normal protein – difference lies in the secondary structure. Normal PrP folds into α-alpha helices, whereas infectious PrP folds into β-sheets.