All metazoan cells examined have a enzyme which deaminates adenine in mRNA to produce inosine which codes as if it were guanine. This editing process changes the genetic message by changing a codon and, hence, an amino acid in a protein. The enzyme that does this binds to double- stranded RNA and is called double-stranded dsRNA adenosine deaminase. In the central nervous system glutamate receptors which are ion channels have proteins which are edited. In one type of editing a glutamate residue in the ion channel is changed to arginine with the result that the influx of calcium ions is blocked. This causes a profound change in the function of the cell, and eliminating this enzyme is associated with acute epilepsy and early death in mice. The serotonin receptor in the central nervous system is also edited, and those changes alter its signal transduction activity. Disruption in the activity of these serotonin receptors are associated with eating disorders and epilepsy. This type of editing has also been found in voltage-gated potassium channels as well as in sialil transferases which add sialic acid to glycoproteins. Editing in that case changes the secretory pathway and half-life of the transferase. There are many unsolved problems associated with the mode of action of this class of editing enzymes. The enzyme only binds to double-stranded RNA formed by a foldback structure involving intron - exon pairing. We propose to solve its three-dimensional structure in order to gain insight into its mode of action. The enzyme is large, containing 1226 amino acids and has several different domains including a catalytic deaminase domain, three double-stranded RNA binding domains and 2 domains which bind with high affinity to left-handed Z-DNA. We propose to solve the structure of some of these isolated domains by X-ray crystallographic or NMR studies in solution. We plan to crystallize larger segments up to and including the entire enzyme. We plan to solve the structure of the enzyme when it is complexed to its substrates double-stranded RNA as well as left-handed DNA. Knowledge of the structure of the enzyme associated with its complex substrates should provide significant insight into the manner in which this enzyme selectively alters the genetic message.
