Chromatographic systems capable of resolving the 2'- and 3'-isomers of valyladenosine and N-acetylvalyladenosine are used to show that valyladenosine isolated from valyl-sRNA is the equilibrium mixture of 2'- and 3'-acyl isomers by the time of chromatography. Acyl migration in N-acetylvalyladenosine is shown to be a rapid, base-catalyzed reaction with a rate 40,000 times the rate of hydrolysis under equivalent conditions. Calculations indicate that the pool of free aminoacyl sRNA exists in vivo as the equilibrium mixture of the 2'- and 3'-aminoacyl isomers
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