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Title: Recognition of, and response to, deaminated bases by archaeal DNA polymerases
Author: Richardson, Tomas Takuyoshi
ISNI:       0000 0004 2730 301X
Awarding Body: University of Newcastle Upon Tyne
Current Institution: University of Newcastle upon Tyne
Date of Award: 2011
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The archaea comprise one of the three domains of life and are often characterised by a propensity for physically- and geochemically-extreme environments. Under such conditions spontaneous DNA deamination events, which ordinarily occur at stochastically insignificant rates, increase in frequency to the point where specialised recognition pathways are required to maintain genomic stability. Archaeal DNA polymerases are unique in their capacity to recognise and respond to deaminated bases, such as uracil and hypoxanthine. For example, the family B DNA polymerases of archaea possess a well-characterised uracil-binding pocket, which, helps prevent replicative bypass of deaminated bases and thus proliferation of fixed mutations. This thesis aims to elucidate additional features of the deaminated base recognition pathways of archaeal DNA polymerases. Here we present studies that concern both the family B and more enigmatic family D DNA polymerases of archaea. Methods employed for investigation of these enzymes include mobility shift assays, targeted mutagenesis, primer extension, exonuclease and uracil-DNA glycosylase assays, as well as time-resolved and steady-state fluorescence analysis. Furthermore, through genetic manipulation of Thermococcus kodakarensis, this work seeks to address previously unanswered questions regarding DNA replication and repair in the archaea. The in vivo studies of deaminated base recognition presented in chapter 5 raise intriguing questions about fundamental aspects of the molecular and cellular biology of archaea.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available