Replication

DNA Replication

What is replication?Unique property of double stranded

molecules of DNASemi-conservative synthesis of DNA based

on a templateDuplication of genetic material The main mechanism that assures

transmission of information from cell to cell and from one generation – to another.

Why replication is required?

Assures exact transmission of information;

Determines self-reproduction; Represents the support of heredity; Assures material linkages between

generation.

When replication takes place?

What are the principles of replication?

Based on template;Complementary;Antiparallel;Two directions;Semi-conservative;Fidel;Very complex.

Bidirectional synthesis

Principles of replication:The monomers (nucleotides) are added to 3’ of

one strand by enzyme DNA polymerase;Newly syntheses strand is complementary to

template strand;Synthesis is semi-conservative: each new

double-stranded molecule contains an old strand (template strand) a new strand (daughter strand).

Semi-conservative mechanism

Components required for replicationsDNA molecule - templateOrigin of replication – point ORIEnzymesNucleotides (dNTP and NTP) SSB proteins

Main enzymes required for replicationDNA-polymerase (I, II,

III – in prokaryotes, a, g, e – in eukaryotes)

Primase DNA-helicases Topoisomerase DNA-ligaseTelomerase

DNA Polymerase

Topoisomerase

What are the main steps of replication?

InitiationElongationTermination

Initiation of replication:Identification of point ori;Formation of replication eye containing two

replication forks:Unfolding of DNADenaturation of DNA

Synthesis of RNA primersInitiation of polymerization of DNA

Point of initiation of replication - ORI

• consists of ~300 bp

• contains sites for interaction with specific proteins required for initiation of replication

ElongationEnlarging of replication eye;Synthesis of DNA in two directions:

Continuous synthesis of leader strand;Interrupted synthesis of lagging strand; synthesis of

Okazaki fragments on lagging strand.

TerminationMeeting of opposite replication forks;RNA primers are removed;Feeling in the gaps;Ligation of all fragments;Renaturation of DNA;

1 mol. DNA 2 mol. DNA.

The replicon – replication unit

• Contains 100-300 kb.

• Contains ORI point

• Contains two replication forks

• In eukaryotic there are numerous replicons

Characteristics of eukaryotic DNA-polymerases DNA polymerase α δ ε β γ

Location Nucleus Nucleus Nucleus Nucleus Mitochondria

Synthetic activityPrimer

extensionSynthesis of

DNADNA repair DNA repair Replication

Additional functions

- Exonuclease Exonuclease - Exonuclease

Peculiarities of replication in eukaryotesIn nucleus replication is carried down by DNA-

polymerases a, dIn mitochondria replication is done by DNA-

polymerase g. Chromosomal DNA contains many points ORI; Replication speed – 20-100 bases/second;Human nuclear genome contains 60000 ORI

points at every 100-150 kb.

Replication of nuclear human DNA is carried out in 6-9 hours

Example:If 1st chromosome will contain 1 ori point,

replication will take place during 15 hours:Contains 260 000 000 bp;Each replication fork will contain 130 000 000

bp Replication speed - 100 bp/sec;

Peculiarities of replication of telomeres

Telomeres:Sequences of nucleotides at the end of chromosomes;(GGGTTAA)n;Form loops;

Telomerase:DNA-polymerase RNA-dependent;Contains snRNA – template for synthesis of telomeric

DNA.

Replication of telomeres

Werner Patient

26 years old

30 years old

Doctor and patient – 30 years old 45 years old