1. Introduction to Molecular Biology
Welcome to Introduction to Bioinformatics
Introduction to Molecular Biology
DNA to protein

2. Molecular biology forms part of a long intellectual tradition
c. 350 BCE - Aristotle begins the Western tradition of natural philosophy
Robert Hook coins the term “cell” in his treatise Micrographia
Anton van Leeuwenhoek discovers bacteria, later spermatozoa
Carl Linnaeus’ Systema Naturae lays the foundations for taxonomy
Charles Darwin’s Origin of the Species
1860s - Louis Pasteur disproves abiogenesis and develops the “Germ theory”
Gregor Mendel describes the “inheritance of traits” in peas.
Ernst Haeckel postulates that the nucleus responsible for heredity
Friedrich Miescher isolates a crude nucleic acid preparation
1880s - Cytologists work out the details of mitosis, meiosis and fertilization

3. Molecular biology forms part of a long intellectual tradition
Walter Sutton proposes a chromosomal theory of heredity
Thomas Hunt Morgan discovers that genes can mutate
1909 – Archibald Garrod proposes the “gene-enzyme” hypothesis
1915 – Morgan and colleagues publish linkage maps of D. melanogaster
1927 – H.J. Muller and L.J. Stadler show that radiation can induce mutation
Fredrick Griffith demonstrates genetic transformation
Barbara McClintock proves genetic recombination – “crossing over”
Avery, MacLeod & McCarty show that DNA carries genetic information
1940s - The “modern synthesis” - E. Mayr, T. Dobzhansky, & J. Huxley
1949 – Erwin Chargaff formulates “Chargaff’s rules” of DNA composition
Hershey and Chase demonstrate that DNA is the genetic material
James Watson and Francis Crick describe the double helix of DNA
1957 – Vernon Ingram - genes determine the sequence of amino acids
Matt Meselson and Frank Stahl prove semiconservative replication

4. Molecular biology forms part of a long intellectual tradition
Arthur Kornberg discovers DNA polymerase
1960 – Sam Weiss and Jerard Hurwitz independently discover RNA polymerase
1960s - Genetic code cracked by Crick, Marshall Nirenberg, Har Gobind Khorana, etc.
Charles Yanofsky & Sydney Brenner show colinearity of DNA & protein
Holley and Zamir determine the structure of a tRNA
20xx – Your contribution!!

5. The “Central Dogma” of Molecular Biology
Term coined by Francis Crick in 1956 to describe the flow
of information in the cell
DNA RNA Protein
Replication
Transcription
Translation

6. Information flow is compartmentalized
DNA
Transcription
Pre-mRNA
Processing
mRNA
Export
mRNA
mRNA
Decay
Translation
Protein
protein
Decay

7. What is the nature of the Gene?
Frederick Griffiths demonstrates “Transformation” of a
heritable character in the bacteria Streptococcus pneumoniae

8. What is the nature of the Gene?
Oswald Avery, Colin MacLeod & Maclyn McCarty
first show that DNA is the “genetic principle”
Enzymes
used to
degrade
proteins

9. The Hershey-Chase experiment confirms
that DNA is the stuff of heredity

10. Erwin Chargaff and his rules
Ratio of nucleotides depends on species
A=T and G=C no matter the organism

11. What is the structure of DNA?
1952
Rosalind Franklin and Maurice Wilkins produce X-ray diffraction images of DNA crystals that suggested that DNA must have some helical arrangement

12. What is the structure of DNA?
1953
Francis Crick and James Watson put together all of the clues and correctly
deduce that DNA is a Double Helix

13. DNA base pairing occurs through hydrogen bonds
A:T pairs: 2 bonds
G:C pairs: 2 bonds

14. The double helix strongly suggested that DNA replication
might proceed by a “semiconservative” process

15. The Meselson-Stahl experiment argues
for strand separation during DNA replication

16. Genes control the amino acid
sequence of proteins
1957 – Vernon Ingram shows that sickle cell haemoglobin varies from wild type by the substitution of one amino acid

17. Genes control the amino acid sequence of proteins
Alteration of amino acid sequence is also
observed in all other hereditary anaemias!

18. DNA cannot directly specify the sequence of amino acids in proteins
Protein synthesis in eukaryotic cells known to take place in the cytoplasm
There must therefore be a SECOND information containing molecule that gets its specificity from DNA, but then moves to the cytoplasm
Attention immediately focuses on RNA – was easy to imagine that it could be produced from a DNA template
Torborn Caspersson and Jean Brachet demonstrated that RNA was mostly in the cytoplasm
Jean Brachet ( )

19. Discovery of mRNA T2 is a bacteriophage that infects E. coli
Completely shuts down normal cellular transcription. Only viral protein is made
T2 RNA always has the same composition as T2 DNA
T2 carries none of its own RNA
32P is incorporated into RNA made after T2 infection
Only about 3-5% of cellular RNA
is messenger RNA!

20. The case for RNA Chemically very similar to DNA Missing methyl
group in uracil
relative to thymine
Hydroxyl group
Chemically very similar to DNA

21. Easy to imagine RNA being produced
from a DNA template

22. There must be a molecular machine that makes RNA from a DNA template
Jerard Hurwitz and Sam Weiss independently discover an enzyme that will only make RNA in the presence of DNA.
The enzyme uses ATP, GTP, CTP, and UTP as precursors
POOR GUYS!
1959 Nobel prize was already awarded to Severo Ochoa
for what turned out later to be the WRONG ENZYME!

23. RNA Polymerase is a molecular machine that carries out transcription

24. RNA is synthesised in the nucleus but travels to the cytoplasm
Cells pulse-labelled with 3H coupled cytidine
T = 15 minutes
T = ~90 minutes
D.M. Prescott

25. Ribosomes are the site of protein synthesis
ribosomes studding the endoplastic reticulum
Shown using radio labelled amino acids in conjunction with ultracentrifugation to isolate
Different cell fractions. Where does the radioactivity end up at various times?

26. Ribosomes and associated rRNAs
are the factories for protein synthesis

27. Crick’s adaptor hypothesis
Can folded RNA act directly as the template for protein synthesis?
Seems unlikely:
the nucleosides chemically want to react with water soluble groups
but many amino acids are polar
no clear way to discriminate chemically similar amino acids
Crick proposes that an adaptor molecule must fit between RNA
and the incoming amino acids, but its nature is unknown
Incoming amino acid
Adaptor molecule
RNA

28. Crick’s adaptors (tRNAs) are themselves
RNA molecules
Self-folding by complementary base pairs gives a structure with several functional domains
Account for ~10% of cellular RNA
abundance
Typically includes several modified, non-standard bases.

29. Zamecnick and Hoagland discover aminoacyl synthetases
Enzymes that added an adaptor (that we now know to be
tRNA) to amino acids prior to their incorporation in proteins
It turns out these tRNAs are Crick’s proposed adaptors
Mahlon Hoagland
Paul Zamecnik

30. Translation proceeds through
a tRNA intermediate

31. Nature of the genetic code
Obvious early on most likely a triplet code in order to code 20 amino acids:
4 x 4 nucleotides can specify 42 = 16 amino acids
4 x 4 x 4 nucleotides can specify 43 = 64 amino acids
Code must be redundant
Not overlapping – Sydney Brenner’s thought experiment
Marshall Nirenberg and Heinrich Matthaei showed that a homopolymer (UUUUUU…. etc. ) produced a poly-phenylalanine protein

32. Khorana's synthetic RNA approach to cracking the genetic code
Example RNA with two repeating units
RNAs with two repeating units:
(UCUCUCU → UCU CUC UCU) produced a polypetide consisting of alternating Serine and CUC codes for Leucine
RNAs with three repeating units:
(UACUACUA → UAC UAC UAC, or ACU ACU ACU, or CUA CUA CUA) produced three different strings of amino acids
RNAs with four repeating units including UAG, UAA, or UGA, produced only dipiptides and tripeptides thus revealing that UAG, UAA and UGA are stop codons.

33. The genetic code is (almost)
universal

34. Amino acids fall into five functional categories

35. Study Question 11 Degeneracy and frequency of amino acids
Most common Leu Gly Ser
Least common Trp Met His

36. Study Question 12 Single mutation from AGA
Silent: |
Hydrophilic/ Hydrophilic: |

37. Study Question 12 Single mutation from AGA
Silent: |
Conservative: |
Hydrophilic/ Hydrophilic: | |
Hydrophilic/ Hydrophobic: |

38. Study Question 12 Single mutation from AGA
Silent: | |
Conservative: |
Hydrophilic/ Hydrophilic: | | |
Hydrophilic/ Hydrophobic: |
Other: |

39. Proteins have four levels of structure

40. The primary structure of proteins is determined by
peptide bonds between amino acids
Formation of this bond is associated with a small +ve DG
Protein synthesis depends on coupled reactions!

41. Secondary structure - the alpha helix
H bonds
Alpha helical conformation is stabilised by hydrogen bonding

42. Secondary structure – beta sheets
Parallel configuration
Antiparallel configuration

43. Secondary structures combine to determine tertiary structure
The enzyme acetylcholinesterase bound to acetylcholine

44. Proteins combine to form quaternary structures
Collagen Haemoglobin

45. Allosteric interactions
+ve
-ve
Interactions (usually with a small molecule) can
alter the shape and activity of an enzyme

46. Enzymes lower the activation energies
associated with biochemical reactions DG
Typical energy of activation is kcal/mol

47. Eukaryotic mRNA must often must be spliced
in order to produce a mature transcript
Exons often correspond to functional protein domains and alternative splicing can give rise to variant proteins