STRUCTURAL PROPERTIES OF FRIEDREICH'S ATAXIA D(GAA) REPEATS

Abstract The expansion of trinucleotide repeat sequences is the underlying cause of a growing number of inherited human disorders. To provide correlations between DNA structure and mechanisms of trinucleotide repeat expansion, we investigated potential secondary structures formed from the complementary strands of d(GAA·TTC) n , a sequence whose expansion is associated with Friedreich’s ataxia. In 50 mM NaCl, pH 7.5, d(GAA) 15 exhibited a cooperative and reversible decrease in large circular dichroism bands at 248 and 272–274 nm over the temperature range of 5–50°C, providing evidence for a base-paired structure at reduced temperatures. Ultraviolet absorbance melting profiles indicated that the melting temperature ( T m ) of d(GAA) 15 was 40°C. At 5°C, the central portion of d(GAA) 15 was hypersensitive to single-strand-specific P1 nuclease degradation and diethyl pyrocarbonate modification, providing evidence for a hairpin conformation. At temperatures between 25 and 35°C in 50 mM NaCl, the triplet repeat region of d(GAA) 15 was uniformly resistant to degradation by P1 nuclease, including the central portion of the sequence. Our results indicate that the structure of d(GAA) 15 is a hairpin at 5°C, unknown but partially base-paired at 37°C, and an approximately random coil above 65°C.