Specificity of double-stranded RNA adenosine deaminases

The specificity of adenosine deaminases that act on RNA (ADARs) is unlike that of most enzymes. ADARs bind their substrate RNA in a sequence-independent manner, and deaminate from <1% to 60% of the adenosines within an RNA at end-reaction. This dissertation describes studies regarding two facets of ADAR specificity. Chapter 2 investigates the role of RNA character in the deamination patterns produced by ADARs. Specifically, this work comprises the first information regarding how RNA secondary structure promotes selective adenosine deamination, with a focus on the function of internal loops within ADAR substrates. These data demonstrate that ADAR1 recognizes internal loops as the end of a helix in a loop size-dependent manner. Chapter 3 compares the specificities of ADAR1 and ADAR2. These data reveal that certain features of the deaminase reaction, such as nearest neighbor preferences, are specific for a given ADAR. These data also suggest that other features of the reaction may be common among the reactions of different ADARs. Comparison of the deamination patterns catalyzed by ADAR1 and ADAR2 hint at a role for binding site size in ADAR specificity. Together, these data substantiate that features of the RNA substrate dictate the number of deamination events that occur within an RNA. Further, these data suggest that features of both the RNA and the enzyme determine how inosine is distributed within an RNA molecule. The work presented herein proposes two models. The first model regards the basis of ADAR recognition of internal loops as effective helix ends. The second model proposes a role for ADAR binding restrictions in ADAR specificity. A unified model of ADAR specificity is described, and experiments aimed at testing these models are proposed.