MEARA sequence repeat of human CstF-64 polyadenylation factor is helical in solution. A spectroscopic and calorimetric study

The primary structure of the human CstF-64 polyadenylation factor contains 12 nearly identical repeats of a consensus motif of five amino acid residue s with the sequence MEAR(A/G). No known function has yet been ascribed to t his motif; however, according to secondary structure prediction algorithms, it should form a helical structure in solution. To validate this theoretic al prediction, we synthesized a 31 amino acid residue peptide (MEARA(6)) co ntaining six repeats of the MEARA sequence and characterized its structure and stability by circular dichroism (CD) spectroscopy and differential scan ning calorimetry (DSC), No effects of concentration on the CD or DSC proper ties of MEARA(6) were observed, indicating that the peptide is monomeric in solution at concentrations up to 2 mM. The far UV-CD spectra of MEARA(6) i ndicates that at a low temperature (1 degrees C) the MEARA(6) peptide has a relatively high helical content (76% at pH 2.0 and 65% at pH 7.0). The eff ects of pH and ionic strength on the CD spectrum of MEARA(6) suggest that a number of electrostatic interactions (e.g., i, i + 3 Arg/Glu ion pair, cha rge-dipole interactions) contribute to the stability of the helical structu re in this peptide. DSC profiles show that the melting of MEARA(6) helix is accompanied by positive change in the enthalpy. To determine thermodynamic parameters of helix-coil transition from DSC profiles for this peptide, we developed a new, semiempirical procedure based on the calculated function for the heat capacity of the coiled state for a broad temperature range. Th e application of this approach to the partial molar heat capacity function for MEARA(6) provides the enthalpy change for helix formation calculated pe r amino acid residue as 3.5 kJ/mol.