The document describes the structure and replication of DNA, highlighting the composition of nucleotides and the double helix model proposed by Watson and Crick. It outlines the various stages of DNA replication, the enzymes involved, and the semiconservative mechanism of replication, which allows for the synthesis of new DNA strands. Additionally, the document touches on the processes of transcription and translation, detailing the roles of RNA and the machinery responsible for protein synthesis.

1. DNA
STRUCTURE &
REPLICATION

2. DNA Structure
DNA is a nucleic acid
The building blocks of DNA are nucleotides, each composed of:
A 5-carbon sugar called deoxyribose
A phosphate group (PO4)
A nitrogenous base
Adenine, Thymine, Cytosine, Guanine

3. The nucleotide structure consists of
the nitrogenous base attached to the 1’
carbon of deoxyribose
the phosphate group attached to the 5’
carbon of deoxyribose
a free hydroxyl group (-OH) at the 3’
carbon of deoxyribose

5. Nucleotides are connected to each other to form a long chain
phosphodiester bond: bond between adjacent nucleotides
 formed between the phosphate group of one nucleotide and the 3’ –OH of
the next nucleotide
The chain of nucleotides has a 5’ to 3’ Orientation

6. 6

7. Determining the 3-dimmensional structure of DNA
involved the work of a few scientists
Erwin Chargaff determined that
amount of adenine = amount of thymine
amount of cytosine = amount of guanine
This is known as Chargaff’s Rules

8. James Watson and Francis Crick, 1953
deduced the structure of DNA using
evidence from Chargaff, Franklin, and
others
proposed a double helix structure

9. The double helix consists of:
2 sugar-phosphate backbones
nitrogenous bases toward the interior of the
molecule
bases form hydrogen bonds with
complementary bases on the opposite
sugar-phosphate backbone

10. The two strands of nucleotides are antiparallel to each other
one is oriented 5’ to 3’, the other 3’ to
5’
The two strands wrap around each other to create the helical shape of
the molecule

12.  In these grooves,
proteins can interact
specifically with
exposed atoms of the
nucleotides (usually by
H bonding) and
thereby recognize and
bind to specific
nucleotide sequences
without disrupting the
base pairing of the
double-helical DNA
molecule

14. A B Z
Shape Broadest Intermedi
ate
Narrowest
Rise per
base pair
2.3 Å 3.4 Å 3.8 Å
Screw
sense
Right
handed
Right
handed
Left handed
Base per
turn
11 10.4 12
Glycosidic
Bond
Anti Anti Alternating Anti
and syn
Comparison of A , B and Z forms of DNA

15. STEPS INVOLVED IN DNA REPLICATION
1. Identification of the origins of replication
2. Unwinding (denaturation) of dsDNA to provide an
ssDNA template
3. Formation of the replication fork; synthesis of RNA
primer
4. Initiation of DNA synthesis and elongation
5. Formation of replication bubbles with ligation of the
newly synthesized DNA segments
6. Reconstitution of chromatin structutre
15

16. DNA replication is the process
where an entire double-stranded
DNA is copied to produce a
second, identical DNA double
helix
Meselson and Stahl concluded
that the mechanism of DNA
replication is the semiconservative
model
Each strand of DNA acts as a
template for the synthesis of a
new strand
16
Q. During which phase
of the cell cycle
does DNA replicate?

17. FACTORS NEEDED FOR REPLICATION
1. DNA HELICASES
to separate the strand
2. GYRASE (Topoisomerases)
unwind the supercoil
3. Single strand binding protein (SSBP)
  activity of helicase
 keep two strand separate
 protect DNA from nuclease degradation
 release after replication
17

18. 4. PRIMOSOME
Complex Protein (contains)
 primase
 helicase (dna B)
 act as primer for DNA synthesis
 removed by polymerase I
18

19. 5. DNA SYNTHESIS CATALYZED BY DNA POLYMERASE
III
 New strand synthesis is catalyzed by DNA poly III
 5'  3' direction
 Antiparallel to parent template
 dNTPs are added one after another to 3‘
 Newly synthesized strand nucleotide is
complementary to parent DNA strand
19

20. 6. DNA LIGASE:
It catalyses the formation of phosphodiester linkage
between small fragment of DNA
7. TERMINATION
 Termination utilization sub (Tus) protein binds to
termination sequence  prevent helicase (dna B)
protein
 Unwinding of DNA helix is stopped
20

21. Initiation of Replication
 Initiation starts at a site called origin of replication
 The origin of replication in E. coli is termed oriC
 origin of Chromosomal replication
 Important DNA sequences in oriC
AT-rich region
DnaA protein binds with the site of origin causing the double
stranded DNA to separate
 The dsDNA is unwound by Helicase (DnaB)
 This generates positive supercoiling ahead of each replication
fork
21

22. Initiation of Replication
 DNA gyrase: relieves tension from the unwinding of
the DNA strands during bacterial replication
 It cuts nicks in both strands of DNA, allowing them to
swivel around one another and then resealing the cut
strands
 The single stranded binding proteins (SSBs) bind to
the exposed bases to prevent them from annealing
Then short (5 to 50 nucleotides) RNA primers are
synthesized by primase
22

23.  These short RNA strands start, or prime DNA synthesis
 DNA polymerases can synthesize DNA only in the 5’ to 3’
direction
 DNA synthesis is semidiscontinuous and bidirectional
 On the leading strand the DNA synthesis is
continuous
On the lagging strand the DNA synthesis is discontinuous
 DNA polymerase III adds complimentary nucleotides
(deoxyribonucleoside triphosphates) in the 5’ to 3’ direction, using
RNA points
 The segments arprimers as starting e called Okazaki fragments
23

24. Building Complimentary
Strands In prokaryotes, there are 3
enzymes known to function in
replication & repair
 DNA polymerase I, II & III
 In eukaryotes, there are 5
enzymes known to function
in replication & repair
DNA pol α, β, γ, δ,ε
24

25. 25

26. Building Complimentary Strands in
prokaryotes
 RNA primers are synthesized by primase and are temporary
 The leading strand (uses 3’-5’ template) is synthesized
continuously
 The lagging strand (uses 5’-3’ template) is synthesized
discontinuously in short fragments
26

27. Building Complimentary Strands in
prokaryotes
DNA polymerase I removes the RNA primers from
the leading strand and fragments from the lagging
strand and replaces them with the appropriate
deoxyribonucleotides
27

28. 28

29. Building Complimentary Strands in
prokaryotes
 DNA ligase joins the Okazaki fragments into one
strand on the lagging strand of DNA through the
formation of a phosphodiester bond
29

30. Replication fork and DNA synthesis30

31. Building Complimentary
Strands in prokaryotes
As the 2 new strands of DNA are
synthesized, 2 double stranded DNA
molecules are produced that automatically
twist into a helix
31

32. PROOF READING
 Fidelity of replication is important is done by DNA Polymerase II
 DNA Polymerase III – beside synthesis also perform proof reading activity
(3’ 5’ exonuclease activity)
 Check during synthesis
 Correction
32

33. PROCESSIVITY
 The average number of nucleotides added before a polymerase
dissociates defines its processivity
 DNA polymerases vary greatly in processivity; some add just a few
nucleotides before dissociating, others add many thousands
33

34. EUKARYOTIC REPLICATION
 Multiple origin site
 Multiple replication bubbles
 Enzymes – 5 types
 DNA polymerase - , , ,  and 
 DNA Poly  = Synthesis of RNA primer (both strand)
Responsible for initiation
 Poly  = Repair of DNA
 Poly  = Replication of mitochondrial DNA
 Poly  = Replication of leading & lagging strand
Proof reading
 Poly  = Lagging strand
34

35. Transcription
 RNA synthesis, or transcription, is the process of transcribing DNA
nucleotide sequence information into RNA sequence information.
 RNA synthesis is catalyzed by a large enzyme called RNA polymerase.
 It takes place in three stages: initiation, elongation, and termination.

36. RNA polymerase performs multiple
functions in transcription
1.It searches DNA for initiation sites, also called promoter sites.
2.It unwinds a short stretch of double-helical DNA to produce a single-
stranded DNA template from which it takes instructions.
3.It selects the correct ribonucleoside triphosphate and catalyzes the
formation of a phosphodiester bond.
4.It detects termination signals that specify where a transcript ends.
5.It interacts with activator and repressor proteins that modulate the rate
of transcription initiation over a wide dynamic range.

37. Transcription cycle
(1) Template binding: RNA polymerase (RNAP) binds to DNA
and locates a promoter (P) melts the two DNA strands to
form a preinitiation complex (PIC).
(2) Chain initiation: RNAP holoenzyme (core + one of multiple
sigma factors) catalyzes the coupling of the first base
(usually ATP or GTP) to a second ribonucleoside
triphosphate to form a dinucleotide.
(3) Chain elongation: The fundamental reaction of RNA
synthesis is the formation of a phosphodiester bond. The 3 -
hydroxyl group of the last nucleotide in the chain
nucleophilically attacks the phosphate group of the incoming
nucleoside triphosphate with the concomitant release of a
pyrophosphate
(4) Chain termination and release: The completed RNA chain
and RNAP are released from the template. The RNAP
holoenzyme re-forms, finds a promoter, and the cycle is
repeated.

38. Transcription cycle

39.  The strand that is transcribed or copied into an RNA
molecule is referred to as the template strand of the
DNA.
 The other DNA strand is frequently referred to as the
coding strand of that gene.

40.  Transcription starts at promoters on the DNA template.
 Promoters are sequences of DNA that direct the RNA
polymerase to the proper initiation site for
transcription.
 Two common motifs are present on the 5 (upstream)
side of the start site.
 They are known as the -10 sequence and the -35
sequence because they are centered at about 10 and
35 nucleotides upstream of the start site.

41. Promoter region of
prokaryotes

42. Transcription process

43. Assembly of Transcription activator
complex in eukaryotes

44. Termination of transcript

45. Processing of primary
transcript

47. Translation
The four letter alphabet of nucleic acid is
translated into entirely different 20 alphabets of
proteins

48. Translation Requirements
 mRNA- Template
 tRNA- Adaptor molecule
 Ribosome- Molecular machinery
 Amino acids- Precursors of protein
 Enzyme and translation factors
 Energy- ATP and GTP

49. mRNA
 Template
 Nucleotides are arranged in triplet
of bases – Codons
 Protein coding region of each
mRNA is composed of a
contiguous, non-overlapping
string of codons called an
opening reading frame (ORF)
 Begins with start codon and end with stop codon

50. tRNA
 Adaptor molecule
 Which link amino acid and mRNA

51. Activation of tRNA
 Some tRNA recognize more than one codon
 Codon-anticodon interaction – Complementary base pairing and
antiparallel
 First two bases of codon and anticodon pair in standard
way
 Codon differ in 1st two bases are read by different tRNA
 Eg. UUA and CUA – Leucine
 1st base of anticodon determine number of codon read by
one tRNA
 C or A – One
 U or G – Two Eg. UUA & UUG and UUU & UUC
 I – three Eg. GCU, GCC, GCA (Ala)

52. rRNA
 Molecular machinery- Protein
Synthesis
 Ribonucleoprotein assemblies
 2/3rd of mass rRNA
 Recognition of start signal –
16SrRNA
 Peptidyl transferase – 23SrRNA

53. Types of rRNA

55. Stages of protein
Synthesis
 Initiation
 Elongation
 Termination

56. Activation
 Activation of amino acids
 Amino-acyl tRNA synthetases, ATP and tRNA
 Form ester with either 3’-OH or 2’-OH of terminal
Adenine residue of tRNA
 Two equivalent ~P used
 Ester linkage and drive the reaction
 Aminoacyl-tRNA synthatases belong to 2
Classess
 Class I acylate at 2’-OH and for Larger and more
hydropobic amino acid
 Class II acylate at 3’-OH (Except Phe-tRNA)

57. Activation

58.  In prokaryotes, Shine-
Dalgarno sequences
 recognized by an initiation
complex
 consisting of a Met amino-
acyl tRNA,
 Initiation Factors (IFs) and
the small ribosomal subunit

59.  GTP hydrolysis by IF2 coincident
with release of the IFs and binding
of the large ribosomal subunit
leads to formation of a complete
ribosome, on the mRNA and ready
to translate.

60.  Nucleotide sequence in mRNA signal where to start
protein synthesis
 The translation of an mRNA begins with codon AUG.
 In eukaryotes cap and in prokaryotes Shine - Dalgarno
sequence tell ribosome where to begin searching for
the start of translation

61. A U G G G C U U AAA G C A G U G C A C G U U
A ribosome on the rough endoplasmic
reticulum attaches to the mRNA molecule.
ribosome

62. A U G G G C U U AAA G C A G U G C A C G U U
It brings an amino acid to the first three
bases (codon) on the mRNA.
Amino acid
tRNA molecule
anticodon
U A C
A transfer RNA molecule arrives.
The three unpaired bases (anticodon)
on the tRNA link up with the codon.

63. A U G G G C U U AAA G C A G U G C A C G U U
Another tRNA molecule comes into
place, bringing a second amino acid.
U A C
Its anticodon links up with the second
codon on the mRNA.

64. A U G G G C U U AAA G C A G U G C A C G U U
A peptide bond forms between the
two amino acids.
Peptide bond

65. A U G G G C U U AAA G C A G U G C A C G U U
The first tRNA molecule releases its amino
acid and moves off into the cytoplasm.

66. A U G G G C U U AAA G C A G U G C A C G U U
The ribosome moves along the mRNA to
the next codon.

67. A U G G G C U U AAA G C A G U G C A C G U U
Another tRNA molecule brings
the next amino acid into place.

68. A U G G G C U U AAA G C A G U G C A C G U U
A peptide bond joins the second
and third amino acids to form a
polypeptide chain.

69. A U G G G C U U AAA G C A G U G C A C G U U
The polypeptide chain gets longer.
The process continues.
This continues until a termination
(stop) codon is reached.
The polypeptide is then complete.

70. Antibiotics inhibiting Translation

71. mRNA
 Contains initiation and termination signal
 Collection of codons – Genetic code
 Degeneracy
 Unambiguous
 Non overlapping
 No punctuation
 Nearly Universal
 Synonyms
 Codons decode the same amino acids
 UUU and UUC – codon for Phe
 AUG – Met and UGG- Trp
 Two (rest), Three (Ile), Four (TGVPA) and Six
Codons (SLR)

72.  XYU and XYC always code same amino acid
 XYA and XYG usually code same amino acid