Intermediacy of Poly(l-proline) II and β-Strand Conformations in Poly(l-lysine) β-Sheet Formation Probed by Temperature-Jump/UV Resonance Raman Spectroscopy

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Abstract
Ultraviolet resonance Raman spectroscopy (UVRR) in combination with a nanosecond temperature jump (T-jump) was used to investigate early steps in the temperature-induced α-helix to β-sheet conformational transition of poly(l-lysine) [poly(K)]. Excitation at 197 nm from a tunable frequency-quadrupled Ti:sapphire laser provided high-quality UVRR spectra, containing multiple conformation-sensitive amide bands. Although un-ionized poly(K) (pH 11.6) is mainly α-helical below 30 °C, there is a detectable fraction (∼15%) of unfolded polypeptide, which is mainly in the poly(l-proline) II (PPII) conformation. However, deviations from the expected amide I and II signals indicate an additional conformation, suggested to be β-strand. Above 30 °C un-ionized poly(K) forms a β-sheet at a rate (minutes) which increases with increasing temperature. A 22−44 °C T-jump is accompanied by prompt amide I and II difference signals suggested to arise from a rapid shift in the PPII/β-strand equilibrium. These signals are superimposed on a subsequently evolving difference spectrum which is characteristic of PPII, although the extent of conversion is low, ∼2% at the 3 μs time limit of the experiment. The rise time of the PPII signals is ∼250 ns, consistent with melting of short α-helical segments. A model is proposed in which the melted PPII segments interconvert with β-strand conformation, whose association through interstrand H-bonding nucleates the formation of β-sheet. The intrinsic propensity for β-strand formation could be a determinant of β-sheet induction time, with implications for the onset of amyloid diseases.