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Title: Structural basis of the Methanothermobacter thermautotrophicus MCM helicase activity
Author: Costa, Alessadro
ISNI:       0000 0001 3388 9799
Awarding Body: Imperial College London (University of London)
Current Institution: Imperial College London
Date of Award: 2007
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The eukaryotic MCM2-7 replicative helicase is a hetero-oligomeric assembly comprising six .homologous polypeptides, which all belong to the superfamily of AAA+ ATPases (ATPases Associated vith various cellular Activities). The MCM complex works as a molecular motor which utilizes the energy produced by ATP hydrolysis to unwind the DNA double helix at the replication fork. Like most archaea, Methanothe.rmobacter thermautotrophicus possesses one copy of the MCM protein which forms a homo-oligomeric assembly and is used as a model for elucidating the mechanism of action of the eukaryotic homologue. Presented here is an electron-microscopy study of this archaeal MCM complex bound to different substrates. Although MCM is characterized by a high degree of polymorphism, I observed that hexameric configurations were.�·stabilized upon DNA binding. I found that the DNA can interact with two distinct regions of the protein complex, either wrapping around the MCM ring, or threading through its central ch'annel. In particular, when treating the protein with a short stretch of DNA, a conformational change occurred within the ATPase domain, Teconfiguring the DNA interacting elements with'in the AAA+ module, which were found projecting inside the central channel of the MCM complex. In contrast, when treating the protein with longer stretches of DNA, the central channel was unoccupied and the DNA was found wrapped around the MCM ring in a configuration similar to that described for the bacterial DnaA initiator factor, an AAA+ ATPase involved in melting of the origin. The latter observation was unexpected and led to the jdef)tificationof a previously unrecognized helixturn- helix DNA binding motif within the outer belt of the MCM complex. Based on these observations I suggested that the MCM protein may have a dual role, contributing to origin melting during the initiation step, and acting as a canonical helicase at the replication fork during the elongation step of DNA replication.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available