Molecular genetics (BIO 16609)
Genetic recombination
• Eka Sunarwidhi Prasedya
•
[email protected]• +6287859972478
• Pusat Unggulan Iptek – Biosains dan Bioteknologi
• https://www.icbb-unram.org/about-pubb-icbb-unram/
Genetic recombination:
1.Homologous Recombination
2. Site-Specific Recombination
3. DNA Transposition
Homologous Recombination at the Molecular
Level
DNA BREAKS ARE COMMON AND INITIATE
RECOMBINATION
Recombination repair DNA breaks by retrieving
sequence information from undamaged DNA
Double-Strand Breaks are Efficiently Repaired
MODELS FOR HOMOLOGOUS
RECOMBINATION
Strand invasion (strand exchange) is a key step in
homologous recombination
Resolving Holliday junctions is a
key step (final step) to finishing
genetic exchange
The double-strand break-repair
model describes many
recombination events
HOMOLOGOUS RECOMBINATION PROTEIN
MACHINES
RecBCD
RecA
RuvAB
RuvC
The RecBCD helicase/nuclease processes broken DNA
molecules for recombination
c-site 5'-GCTGGTGG-3'.
Structure of RecBCD
Chi sites control RecBCD (GCTGGTGG)
RecA protein assembles on single-stranded DNA and
promotes strand invasion
Three views of the RecA filament
Polarity of RecA assembly
Newly based-paired partners are established within RecA
filament
RecA homologs are present in all organism
Human Rad51 RecA
Rad A of Archaea
The RuvAB complex specifically recognizes Holliday
junctions and promote branch migration
Branch migration can either enlarge heteroduplex regions or
release newly synthesized DNA as a single strand
Structure of RuvA and model of RuvAB bound to Holliday junction
Enzyme-catalyzed double branch migration at a Holliday junction.
In E. coli, a tetramer of the RuvA protein (green) and two hexamers of the RuvB
protein (pale gray) bind to the open form of the junction. The RuvB protein uses the
energy of ATP hydrolysis to move the crossover point rapidly along the paired DNA
helices, extending the heteroduplex region as shown. There is evidence that similar
proteins perform this function in vertebrate cells. (Image courtesy of P. Artymiuk;
modified from S.C. West, Cell 94:699–701, 1998.)
RuvC cleaves specific strands at the Holliday junction to
finish recombination
Structure of RuvC and model of RuvC dimer bound to Holliday
junction
RecBCD
RecA
RuvAB
RuvC
How to resolve a recombination intermediate with two Holliday
junctions
HOMOLOGOUS
RECOMBINATION IN
EUKARYOTES
Homologous recombination has
additional functions in eukaryotes
It is required to pair homologous
chromosomes in preparation for the
first nuclear division and for
segregation during meiosis
Meiotic recombination between
homologous chromatids
Meiotic recombination also frequently gives rise to crossing over
between genes on the two homologous parental chromosomes
Programmed generation of double-stranded DNA breaks
occurs during meiosis
MRX protein processes the
cleaved DNA ends for
assembly of the RecA-like
strand exchange proteins
(DMC1, Rad51)
DMC1 specifically functions
in meiotic recombination
Many proteins function together to promote meiotic
recombination (Rad51, DMC1, Rad51 paralogs, Rad52,
Rad54)
Site-Specific recombination & Transposition
Conservative site-specific recombination
Three types of CSSR
Structures involved in CSSR
Recombination by a serine
recombinase
Recombination by a tyrosine
recombinase
Conservative site-specific recombination
was discovered in bacteriophage lambda
The insertion of a circular bacteriophage
lambda DNA chromosome into the
bacterial chromosome.
The life cycle of bacteriophage
lambda.
The double-stranded DNA lambda
genome contains 50,000 nucleotide
pairs and encodes 50–60 different
proteins. When the lambda DNA
enters the cell, the ends join to
form a circular DNA molecule.
This bacteriophage can multiply in
E. coli by a lytic pathway, which
destroys the cell, or it can enter a
latent prophage state. Damage to a
cell carrying a lambda prophage
induces the prophage to exit from
the host chromosome and shift to
lytic growth (green arrows). Both
the entrance of the lambda DNA to,
and its exit from, the bacterial
chromosome are accomplished by
a conservative site-specific
recombination event, catalyzed by
the lambda integrase enzyme (see
Figure 5–80).
DNA inversion by the Hin recombinase of Salmonella
Transposition
Some genetic elements move to new chromosomal
locations by transposition
Genes make up only a small portion of the eukaryotic
chromosomal DNA
Representation of the nucleotide sequence content of the human genome.
LINES, SINES, retroviral-like elements, and DNA-only transposons are all mobile genetic elements that
have multiplied in our genome by replicating themselves and inserting the new copies in different positions.
Simple sequence repeats are short nucleotide sequences (less than 14 nucleotide pairs) that are repeated again
and again for long stretches. Segmental duplications are large blocks of the genome (1000–200,000
nucleotide pairs) that are present at two or more locations in the genome. Over half of the unique sequence
consists of genes and the remainder is probably regulatory DNA. Most of the DNA present in
heterochromatin, a specialized type of chromatin that contains relatively few genes, has not yet been
sequenced
The cut-and-paste
mechanism of transposition
(DNA transposons)
Replicative transposition
(DNA transposons)
Virus-like
retrotransposons
Some virus use a transposition mechanism to move themselves
into host cell chromosomes
Reverse transcriptase
Poly-A retrotransposon
The Nobel Prize in Physiology or Medicine 1983
"for her discovery of mobile genetic elements"
Barbara McClintock
USA
Cold Spring Harbor Laboratory
Cold Spring Harbor, NY, USA
b. 1902
d. 1992
