slide1 n.
Skip this Video
Download Presentation
Part 3 Genetic Information Transfer

Loading in 2 Seconds...

play fullscreen
1 / 115

Part 3 Genetic Information Transfer - PowerPoint PPT Presentation

  • Uploaded on

Part 3 Genetic Information Transfer. The biochemistry and molecular biology department of CMU. Central dogma. replication. transcription. translation. DNA. protein. RNA. reverse transcription. Replication: synthesis of daughter DNA from parental DNA

I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
Download Presentation

PowerPoint Slideshow about 'Part 3 Genetic Information Transfer' - bijan

Download Now An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.

- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript

Part 3

Genetic Information Transfer

The biochemistry and molecular biology department of CMU


Central dogma







reverse transcription

Replication: synthesis of daughter DNA from parental DNA
  • Transcription: synthesis of RNA using DNA as the template
  • Translation: protein synthesis using mRNA molecules as the template
  • Reverse transcription: synthesis of DNA using RNA as the template
Chapter 10

DNA Replication



parental DNA

DNA replication

  • A reaction in which daughter DNAs are synthesized using the parental DNAs as the template.
  • Transferring the genetic information to the descendant generation with a high fidelity

daughter DNA


Daughter strand synthesis

  • Chemical formulation:
  • The nature of DNA replication is a series of 3´- 5´phosphodiester bond formation catalyzed by a group of enzymes.

DNA replication system

Template: double stranded DNA


Primer: short RNA fragment with a free 3´-OH end

Enzyme:DNA-dependent DNA polymerase (DDDP),

other enzymes,

protein factor

characteristics of replication
Characteristics of replication
  • Semi-conservative replication
  • Bidirectional replication
  • Semi-continuous replication
  • High fidelity

Semiconservative replication

Half of the parental DNA molecule is conserved in each new double helix, paired with a newly synthesized complementary strand. This is called semiconservative replication



The genetic information is ensured to be transferred from one generation to the next generation with a high fidelity.


Replication starts from unwinding the dsDNA at a particular point (called origin), followed by the synthesis on each strand.

The parental dsDNA and two newly formed dsDNA form a Y-shape structure called replication fork.

§1.2 Bidirectional Replication

Bidirectional replication

Once the dsDNA is opened at the origin, two replication forks are formed spontaneously.

These two replication forks move in opposite directions as the syntheses continue.


Replication of prokaryotes

The replication process starts from the origin, and proceeds in two opposite directions. It is named  replication.


Chromosomes of eukaryotes have multiple origins.

The space between two adjacent origins is called the replicon, a functional unit of replication.

Replication of eukaryotes


The daughter strands on two template strands are synthesized differently since the replication process obeys the principle that DNA is synthesized from the 5´ end to the 3´end.

§1.3 Semi-continuous Replication


Leading strand

On the template having the 3´- end, the daughter strand is synthesized continuously in the 5’-3’ direction. This strand is referred to as the leading strand.


Okazaki fragments

Many DNA fragments are synthesized sequentially on the DNA template strand having the 5´- end. These DNA fragments are called Okazaki fragments. They are 1000 – 2000 nt long for prokaryotes and 100-150 nt long for eukaryotes.

The daughter strand consisting of Okazaki fragments is called thelagging strand.


Semi-continuous replication

Continuous synthesis of the leading strand and discontinuous synthesis of the lagging strand represent a unique feature of DNA replication. It is referred to as thesemi-continuous replication.


§2.1 DNA Polymerase

DNA-pol of prokaryotes

  • The first DNA- dependent DNA polymerase (short for DNA-pol I) was discovered in 1958 by Arthur Kornberg who received Nobel Prize in physiology or medicine in 1959.
Later, DNA-pol II and DNA-pol III were identified in experiments using mutated E.coli cell line.
  • All of them possess the following biological activity.

1. 53 polymerizing

2. exonuclease


DNA-pol I

  • Mainly responsible for proofreading and filling the gaps, repairing DNA damage
klenow fragment

N end

DNA-pol Ⅰ

C end


Klenow fragment
  • small fragment (323 AA): having 5´→3´ exonuclease activity
  • large fragment (604 AA): called Klenow fragment, having DNA polymerization and 3´→5´exonuclease activity

DNA-pol II

  • Temporary functional when DNA-pol I and DNA-pol III are not functional
  • Still capable for doing synthesis on the damaged template
  • Participating in DNA repairing


  • A heterodimer enzyme composed of ten different subunits
  • Having the highest polymerization activity (105 nt/min)
  • The true enzyme responsible for the elongation process
structure of dna pol iii
Structure of DNA-pol III

α: has5´→ 3´ polymerizing activity

ε:has3´→ 5´ exonuclease activity and plays a key role to ensure the replication fidelity.

θ: maintain heterodimer structure


DNA-pol of eukaryotes

DNA-pol : initiate replication and synthesize primers



DNA-pol : replication with low fidelity


DNA-pol : polymerization in mitochondria

DNA-pol : elongation


DNA-pol : proofreading and filling gap

DNA-pol I


Also called DnaG

Primase is ableto synthesize primers using free NTPs as the substrate and the ssDNA as the template.

Primers are short RNA fragments of a several decades of nucleotides long.

§2.2 Primase


Primers provide free 3´-OH groups to react with the -P atom of dNTP to form phosphoester bonds.

Primase, DnaB, DnaC and an origin form a primosomecomplex at the initiation phase.

§2.3 Helicase

Also referred to as DnaB.

It opens the double strand DNA with consuming ATP.

The opening process with the assistance of DnaA and DnaC


§2.4 SSB protein

  • Stand for single strand DNA binding protein
  • SSB protein maintains the DNA template in the single strand form in order to
    • prevent the dsDNA formation;
    • protect the vulnerable ssDNA from nucleases.

Opening the dsDNA will create supercoil ahead of replication forks.

The supercoil constraint needs to be released by topoisomerases.

§2.5 Topoisomerase


The interconversion of topoisomers of dsDNA is catalyzed by a topoisomerase in a three-step process:

    • Cleavage of one or both strands of DNA
    • Passage of a segment of DNA through this break
    • Resealing of the DNA break

Topoisomerase I (topo I)

Also called -protein in prokaryotes.

It cuts a phosphoester bond on one DNA strand, rotates the broken DNAfreely around the other strand to relax the constraint,and reseals the cut.


Topoisomerase II (topo II)

It is named gyrase in prokaryotes.

It cuts phosphoester bonds on both strands of dsDNA, releases the supercoil constraint, and reforms the phosphoester bonds.

It can change dsDNA into the negative supercoil state with consumption of ATP.


Connect two adjacent ssDNA strands by joining the 3´-OH of one DNA strand to the 5´-P of another DNA strand.

Sealing the nick in the process of replication, repairing, recombination, and splicing.


§2.7 Replication Fidelity

  • Replication based on the principle of base pairing is crucial to the high accuracy of the genetic information transfer.
  • Enzymes use two mechanisms to ensure the replication fidelity.
    • Proofreading and real-time correction
    • Base selection

Proofreading and correction

  • DNA-pol I has the function to correct the mismatched nucleotides.
  • It identifies the mismatched nucleotide, removes it using the 3´- 5´ exonuclease activity, add a correct base, and continues the replication.

Exonuclease functions

5´→3´ exonuclease activity

cut primer or excise mutated segment

3´→5´ exonuclease activity

excise mismatched nuleotides

sequential actions
Sequential actions
  • Initiation: recognize the starting point, separate dsDNA, primer synthesis, …
  • Elongation: add dNTPs to the existing strand, form phosphoester bonds, correct the mismatch bases, extending the DNA strand, …
  • Termination: stop the replication

§3.1 Replication of prokaryotes

a. Initiation

  • The replication starts at a particular point called origin.
  • The origin of E. coli, oriC, is at the location of 82.
  • The structure of the origin is 248 bp long and AT-rich.

Structure of ori C

  • Three 13 bp consensus sequences
  • Two pairs of anti-consensus repeats

Formation of replication fork

  • DnaA recognizes ori C.
  • DnaB and DnaC join the DNA-DnaA complex, open the local AT-rich region, and move on the template downstream further to separate enough space.
  • DnaA is replaced gradually.
  • SSB protein binds the complex to stabilize ssDNA.

Primer synthesis

  • Primase joins and forms a complex called primosome.
  • Primase starts the synthesis of primers on the ssDNA template using NTP as the substrates in the 5´- 3´ direction at the expense of ATP.
  • The short RNA fragments provide free 3´-OH groups for DNA elongation.

Releasing supercoil constraint

  • The supercoil constraints are generated ahead of the replication forks.
  • Topoisomerase binds to the dsDNA region just before the replication forks to release the supercoil constraint.
  • The negatively supercoiled DNA serves as a better template than the positively supercoiled DNA.

b. Elongation

  • dNTPs are continuously connected to the primer or the nascent DNA chain by DNA-pol III.
  • The core enzymes (、、and ) catalyze the synthesis of leading and lagging strands, respectively.
  • The nature of the chain elongation is the series formation of the phosphodiester bonds.

The synthesis direction of the leading strand is the same as that of the replication fork.

  • The synthesis direction of the latest Okazaki fragment is also the same as that of the replication fork.

Lagging strand synthesis

  • Primers on Okazaki fragments are digested by RNase.
  • The gaps are filled by DNA-pol I in the 5´→3´direction.
  • The nick between the 5´end of one fragment and the 3´end of the next fragment issealed by ligase.

c. Termination

  • The replication of E. coli is bidirectional from one origin, and thetwo replication forksmust meet at one point called ter at 32.
  • All the primers will be removed, and all the fragments will be connected by DNA-pol I and ligase.

§3.2 Replication of Eukaryotes

  • DNA replication is closely related with cell cycle.
  • Multiple origins on one chromosome, and replications are activated in a sequential order rather than simultaneously.


  • The eukaryotic origins are shorter than that of E. coli.
  • Requires DNA-pol  (primase activity) and DNA-pol  (polymerase activity and helicase activity).
  • Needs topoisomerase and replication factors (RF) to assist.

b. Elongation

  • DNA replication and nucleosome assembling occur simultaneously.
  • Overall replication speed is compatible with that of prokaryotes.


  • The terminal structure of eukaryotic DNA of chromosomes is called telomere.
  • Telomere is composed of terminal DNA sequence and protein.
  • The sequence of typical telomeres is rich in T and G.
  • The telomere structure is crucial to keep the termini of chromosomes in the cell from becoming entangled and sticking to each other.


  • The eukaryotic cells use telomerase to maintain the integrity of DNA telomere.
  • The telomerase is composed of

telomerase RNA

telomerase association protein

telomerase reverse transcriptase

  • It is able to synthesize DNA using RNA as the template.

Significance of Telomerase

  • Telomerase may play important roles is cancer cell biology and in cell aging.

§4.1 Reverse Transcription

The genetic information carrier of some biological systems is ssRNA instead of dsDNA (such as ssRNA viruses).

The information flow is from RNA to DNA, opposite to the normal process.

This special replication mode is called reverse transcription.


Reverse transcription

Reverse transcription is a process in which ssRNA is used as the template to synthesize dsDNA.


Process of Reverse transcription

Synthesisof ssDNA complementary to ssRNA, forming a RNA-DNA hybrid.

Hydrolysis ofssRNA in the RNA-DNA hybrid by RNase activity of reverse transcriptase, leaving ssDNA.

Synthesis of the second ssDNA using the left ssDNA as the template, forming a DNA-DNA duplex.


Reverse transcriptase

Reverse transcriptase is the enzyme for the reverse transcription. It has activity of three kinds of enzymes:

RNA-dependent DNA polymerase


DNA-dependent DNA polymerase


Significance of RT

An important discovery in life science and molecular biology

RNA plays a key role just like DNA in the genetic information transfer and gene expression process.

RNA could be the molecule developed earlier than DNA in evolution.

RT is the supplementary to the central dogma.


Significance of RT

This discovery enriches the understanding about the cancer-causing theory of viruses. (cancer genes in RT viruses, and HIV having RT function)

Reverse transcriptase has become a extremely important tool in molecular biology to select the target genes.


§5.1 Mutation

Mutation is a change of nucleic acids in genomic DNA of an organism. The mutation could occur in the replication process as well as in other steps of life process.

consequences of mutation
Consequences of mutation
  • To create a diversity of the biological world; a natural evolution of biological systems
  • To lead to the functional alternation of biomolecules, death of cells or tissues, and some diseases as well
  • Changes of genotype, but no effect on phenotype

Mutation caused by chemicals

  • Carcinogens can cause mutation.
  • Carcinogens include:
    • Food additives and food preservatives; spoiled food
    • Pollutants: automobile emission; chemical wastes
    • Chemicals: pesticides; alkyl derivatives; -NH2OH containing materials

§5.3 Types of Mutation

a. Point mutation (mismatch)

Point mutationis referred to as the singlenucleotide alternation.

Transition:the base alternation from purine to purine, or from pyrimidine to pyrimidine.

Transversion: the base alternation between purine and pyrimidine, and vise versa.


Hb mutation causing anemia

Singlebase mutation leads to one AA change, causing disease.


b.Deletion and insertion

Deletion: one or more nucleotides aredeleted from the DNA sequence.

Insertion: one or more nucleotides are inserted into the DNA sequence.

Deletion and insertion can cause the reading frame shifted.

frame shift mutation
Frame-shift mutation



Ala Val His Val

Deletion C


Glu Tyr Met Ser



It is an exchange of large DNA fragments. It can be either reverse the direction or recombination between chromosomes.

1. Site-specific recombination

2. Homologous genetic recombination

3. DNA transposition


§5.4 DNA Repairing

  • DNA repairing is a kind response made by cells after DNA damage occurs, which may resume their natural structures and normal biological functions.
  • DNA repairing is a supplementary to the proofreading-correction mechanism in DNA replication.

Light repairing



Excision repairing

One of the most important and effective repairing approach.

UvrA and UvrB: recognize and bind the damaged region of DNA.

UvrC: excise the damaged segment.

DNA-pol Ⅰ: synthesize the DNA segment to fill the gap.

DNA ligase: seal the nick.


Xeroderma pigmentosis (XP)

XP is an autosomal recessive genetic disease. Patients will be suffered with hyper-sensitivity to UV which results in multiple skin cancers.

The cause is due to the low enzymatic activity for the nucleotide excision-repairing process, particular thymine dimer.

excision repairing
Excision repairing

DNA polymeraseⅠ


DNA ligase


Recombination repairing

  • It is used for repairing when a large segment of DNA is damaged.
  • Recombination protein RecA, RecB and RecC participate in this repairing.

SOS repairing

  • It is responsible for the situation that DNA is severely damaged and the replication is hard to continue.
  • If workable, the cell could be survived, but may leave many errors.
  • In E. coli, uvr gene and rec gene as well as Lex A protein constitute a regulatory network.