L07 Chem 481 Lecture Notes - Lecture 10: Holliday Junction, Homologous Chromosome, Genetic Recombination
13 Dec 2016
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21 September 2016
Lecture 10: Homologous Recombination
I. DNA Restructuring and Recombination
A. Overview
1. Major processes include (all simultaneous in real life)
a. Restriction and modification
• Often associated with bacterial defense and powerful genome modification via
restriction
b. Recombination of DNA
c. A normal cycle of damage and repair
d. Gene transposition and amplification
• Amplification and duplication are difficult to study
• Many cells duplicate certain genes (typically terminally differentiated cells)
2. Genetic information is shuffled by recombination
a. Genetic recombination rearranges genetic information, creating new associations
b. Recombination involving similar DNA sequences is called homologous recombination
• Programmed design for this, but also involves some repair mechanisms
c. The process underlying homologous recombination is termed general recombination
• This requires the breakage and reunion of DNA strands
d. Recombination involving very different nucleotide sequences is non-homologous
recombination
e. Transposition is the enzymatic insertion of a transposon, a mobile segment of DNA
f. Non-homologous and illegitimate recombination are the same thing
B. Recombination Models
1. The Holliday Model for Homologous Recombination
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a. Two homologous DNA duplexes first juxtapose so that their sequences are aligned, a
process of chromosome pairing called synapsis
• Illustrated with strands in same direction close to each other
b. Recombination starts with introduction of small nicks at homologous sites on the two
chromosomes
c. Duplexes partially unwind, and the free, single-stranded end of one duplex begins to base
pair with its nearly complementary single-stranded region along the intact strand in the
other duplex
• This process is called strand invasion
• Key process in homology because it allows annealing to the homologous duplex
(after both strands nicked and unwound slightly)
d. Ligation follows forming a Holliday junction
• The crossover point = a Holliday junction (facilitated by protein)
• The junction is then resolved and migrates with the help of special machinery
e. Isomerization can occur to form duplexes in different orientation
• Then the strands are cut and rejoined
• This can form either non-recombinant “patch-spliced” hetero-duplex
• Or it can form a recombinant hetero-duplex if cut in the orientation to exchange more
genetic information
2. The Holliday Junction (synthetic visualization)
a. Highly complementary sequences allow these duplexes to form
b. Only small disturbances in base pairing occur and it doesn’t require much bending or
twisting of DNA to form
3. Holiday Model Geometry
a. If is important to look at the polarity of each strand, how the similar directions will be the
interacting parts of the duplex
b. Only strands of like polarity exchange DNA during recombination
c. One crossover event is enough if close enough to the telomeric site (near the end of the
linear chromosome)
d. Can often exchange in the middle of the chromosome, but usually involves at least two
events
4. The Untwisted Holliday Junction
a. Can have cuts EW (horizontal) or NS (vertical) which results in different strand
orientations for re-annealing
b. The directionality of cutting appeals random in the cell, can’t tell the difference usually
5. Messelson-Radding Model of Homologous Recombination
a. Since we haven’t yet found a nuclease to provide the double nicking suggested by the
Holliday model, the Messelson model requires only 1 nick (more likely)
b. 1 strand is cut and there is a displacement o fa single strand, which invades the
homologous DNA to form a D-loop
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