~(13)C Shifts Reveal a Clear Pattern along Strands in β Hairpins

摘要

It has long been recognized that ~(13)C, ~(13)C_α and ~(13)C_β chemical shift deviations (CSDs) from random coil values strongly correlate with protein backbone conformations [1,2]. ~(13)C, ~(13)Cα CSDs are generally positive (more downfield relative to random coil shifts) in a-helices and negative (more upfield) in β-sheets, while ~(13)C_β have opposite trend in the two types of secondary structures. Although the range of ~(13)C CSDs occurring in (3-sheets slightly overlaps with the narrower range for a-helices, ~(13)C chemical shifts are very useful for assigning secondary structure and thereby determining the 3D structure of a protein [3]. Since both backbone H_α and amide protons Hn of (3-strands display an i/i+2 periodicity of downfield shifts in a (3-hairpin or at the edge of a (3-sheet, only the more positive CSDs are used for estimating hairpin fold population [4,5]. On the other hand, ~(13)C_α and ~(13)C_β CSDs were also found to be a potential means to quantify P-hairpin fold population [6,7]. The CSDs of ~(13)C_α and ~(13)Cp were separately averaged for all the strand residues in the considered peptide, excluding the non-protected N- and C-terminal residues, to estimate hairpin fold population. This implies that detailed pattern for ~(13)C CSDs along the (3-strands do not appear to have been recognized in literature yet, unlike the case of 'H_α and ~1H_N shifts. In our aromatic-free hairpin model, we observed the diagnostic ~(13)C, ~(13)C_α and ~(13)C_β shifts for β-structuring are, almost exclusively, associated with the cross-strand hydrogen-bonded residues. Their CSD magnitudes correlate qualitatively well with haiipin fold population based on measurements under different temperatures and incorporating different turn sequences into the hairpin model. HFIP (hexafluoroisopropanol) is shown to greatly influence ~(13)C shifts, and the random coil values applied to derive the (3-strand CSD partem should be adjusted when this co-solvent is present.