Topoisomerase (topos)

Question 1

topoisomerases =

Answer 1

enzymes that alter the topological state of DNA in cells

e.g. interchange between relaxed and supercoiled DNA in bacteria

Question 2

on closed circular DNA…

Answer 2

• DNA pol moving along causes unwinding and some overwinding causes the formation of positive supercoils ahead of the polymerase
• DNA gyrase = works mostly ahead of the replication fork - relaxes the positive supercoils and can introduce negative supercoils
• Topo IV = works mostly behind the replication fork - decatenation of circles of DNA = relax into separate daughter circles + can also be used to relax positive supercoils ahead of the replication fork

Question 3

DNA gyrase and Topo IV are both type II topoisomerases:

Answer 3

• are large tetrameric complexes
• they cleave both strands of the DNA, in contrast to other type I topos that cleave only one strand
• in all cases, the enzyme is linked to the DNA by the formation of a phosphodiester bond between an active site tyrosine and a phosphate group on the DNA backbone
• all organisms have topoisomerases, but only DNA gyrase can perform the energetically unfavourable intersection of supercoils into DNA

Question 4

Topoisomerase II cycle:

Answer 4

1) a section of DNA (the G-segment gate) is bound
2) subsequent binding of ATP closes the top gate, trapping the second piece of DNA (the T-segment, transferred)
3) the G-segment is then cleaved, forming a gate
4) the ends of the DNA are ‘held’ via a covalent bond to Tyr

Question 5

differences gyrase and topo IV:

Answer 5

• gyrase and topo IV both have beta-pinwheel arrangements in their structure and can bend DNA around themselves - however, topo IV cannot put in supercoils = must have a different arrangement
• gyrase has a highly conserved sequence termed the ‘GyrA box’ - in gyrase mutants in which this sequence is deleted cannot wrap DNA or introduce supercoils