Three major classes of transposable elements
DNA only transposons: moves as DNA, either by cut and paste or replicative pathways
Retroviral-like retrotransposons: moves via an RNA intermediate
nonretroviral retrotransposons: moves via an RNA ntermediate that is often produced from a neighboring promoter
What is the mechanism by which more than 50% of our genome has been invaded by genetic elements?
gain/ loss of information--> genome gets bigger or smaller
conservative: on end, there's a conserved sequence for DNA to be inserted at that point
insertion can occur almost anywhere; no specificity
What is common about the three major classes of transposable elements?
- short DNA fragments that are recognized only by the transposase encoded by that element
- on the end is a short repeat of nucleotide that is recognized by enzymes, either transposases or integrases
Explain mobile genetic elements
- They are short fragments that can bring other elements
- They have to bring integrase/ transposase genes; have to have a little sequence at the end
What is a conservative site specific example ?
lambda phage: virus that infects bacteria
- can do three things:
- - can integrate self into host genome (integrase latches on in site-specific way, cuts them, and integrates lambda DNA into host
- the same integrase that inserted it can remove; proteins are produced; phage DNA leaves and gets packaged. Lysis/ lytic pathway
an enzyme that conducts at least some of the DNA breakage and joining reactions needed for the element to move
After the recombination process of lambda causes the DNA protein complex to bind to the bacterial chromosome adn bring the bacterial and pahge chromosome together, what happens?
the integrase catalyzes the required cutting and resealing reactions that result in recombination
Because a short region of sequence homology in the two joined sequences, a tiny heteroduplex joint is formed that is seven nucleotide pairs long
Explaint he insertion of a circular phage DNA chromosome into the bacterial chromosome.
The lambda integrase enzyme binds to a specific attachment-site DNA sequence on each chromosome, where it makes cuts that bracket a short homologous DNA sequence
The integrase then switches the partner strands and reseals them to form a heteroduplx joint that is 7 nucleotide pairs long.
A total of four strand-breaking and strand-joining reactions are required; for each of them, the energy of the cleaved phosphodiester bond is stored in a transient covalent linkage between the DNA and teh enzyme, so that DNA strand resealing occurs without a requirement for ATP or DNA ligase
What is the life cycle of the HIV retrovirus?
The retrovirus genome consists of an RNA molecule that is 8500 nucleotides; two such molecules are typicalyl packaged into each viral particle.
The enzyme reverse transcriptase makes a DNA copy of the viral RNA molecule and then a second DNA strand, generating a double-stranded DNA copy of the RNA genome
The integration of this DNA double helix into the host chromosome is then catalyzed by a virus-encoded integrase enzyme
This integration is required for synthesis of new viral RNA molecules by the host cell RNA polymerase, the enzyme that transcribes DNA into RNA
Structure of reverse transcriptase
- thumbs and fingers taht close around the substrate (RNA), creating a complementary strand of DNA
- contains RNAse, which cuts RNA and chews it up
- It is an RNA dependent DNA polymerase
its active site degrades RNA strands; the H signifies it only works on ds RNA
What can reverse transcriptase do?
it can flip around and make the ds DNA
integrated into host chromosome
Integrase creates what?
it creates sticky ends; it cuts target chromosome and integrates it
not reversible; once in, its in
Explain transposition by either a retrovirus or a retroviral like retrotransposon
- 1) A dsDNA is produced by reverse transcriptase
- 2) In an initial step, the integrase enzyme forms a DNA loop and cuts one strand at each end of the viral DNA sequence, exposing new 3'-OH groups.
- 3) Each of these 3'-OH ends then directly attacks a phosphodiester bond on opposite strands of a randomly selected site on a target chromosome.
- 4) This inserts the viral DNA sequence into the target chromosome, leaving short gaps on each side that are filled in by DNA repair processes
- 5) Because of the gap filling, this mechanism leaves short repeats of target DNA sequence on each side of the integrated DNA segment
Explain Cre recombinase.
- It can insert or remove
- It recognizes lox sites that are recognized by pre recombinase
Animal lacks Cre recombinase--> injected in
Which two terms are synonymous?
recombinase and integrase