Ionized Trilysine: A Model System for Understanding the Nonrandom Structure of Poly-l-lysine and Lysine-Containing Motifs in Proteins.
ABSTRACT It is now well-established that different amino acid residues can exhibit different conformational distributions in the unfolded state of peptides and proteins. These conformational propensities can be modulated by nearest neighbors. In the current study, we combined vibrational and NMR spectroscopy to determine the conformational distributions of the central and C-terminal residues in trilysine peptides in aqueous solution. The study was motivated by earlier observations suggesting that interactions between ionized nearest neighbor residues can substantially change conformational propensities. We found that the central lysine residue predominantly adopts conformations that are located at the upper border of the upper left quadrant of the Ramachandran plot and the left border of the polyproline II region. We term this type of conformation deformed polyproline II (pPII(d)). The structures of less populated subensembles of trilysine resemble are comparable with structures at the i + 1 position of type I and type II β-turns. For the C-terminal residue, however, we obtained a mixture of polyproline II, β-strand, and right-handed helical conformations, which is typical for lysine residues in alanine- and glycine-based peptides. Our data thus indicate that the terminal lysines modify and restrict the conformational distribution of the central lysine residue. DFT calculations for ionized trilysine and lysyllysyllysylglycine in vacuo indicate that the pPII(d) is stabilized by a rather strong hydrogen bond between the NH(3)(+) group of the central lysine and the carbonyl group of the C-terminal peptide. This intramolecular hydrogen bonding induces optical activity in the C-terminal CO stretching vibration, which leads to an unusual and relatively intense positive Cotton band. Additionally, we analyzed the amide I' band profile of ionized triornithine in water. Ornithine is structurally similar to lysine in that its side chain is terminated with an amino group; however, the side chain of ornithine is shorter than lysine's side chain by one methylene group. We found that the conformational distribution of the central ornithine in this peptide must be very similar to that of the central lysine residue in trilysine. This suggests that the ionized ammonium group, which lysine and ornithine side chains have in common, is the main determinant of their conformational propensities at the central position in the respective tripeptides. The results of a DFT-based geometry optimization confirm this notion. In principle, our results suggest that lysine-rich segments in unfolded/disordered proteins and peptides can switch between different types of local order, i.e., an extended pPII(d)-like conformation and transient turns. However, for longer polylysine segments nonlocal interactions between side chains might impede the formation of turns, thus enabling the formation of pPII(d)-helix segments.
Article: The pH-Independence of Trialanine and the Effects of Termini Blocking in Short Peptides: A Combined Vibrational, NMR, UVCD, and Molecular Dynamics Study.[show abstract] [hide abstract]
ABSTRACT: Several lines of evidence now well establish that unfolded peptides in general, and alanine in specific, have an intrinsic preference for the polyproline II (pPII) conformation. Investigation of local order in the unfolded state is, however, complicated by experimental limitations and the inherent dynamics of the system, which has in some cases yielded inconsistent results from different types of experiments. One method of studying these systems is the use of short model peptides, and specifically short alanine peptides, known for predominantly sampling pPII structure in aqueous solution. Recently, He et al. (J. Am. Chem. Soc. 2012, 134, 1571-1576) proposed that unblocked tripeptides may not be suitable models for studying conformational propensities in unfolded peptides due to the presence of end effect, i.e. electrostatic interactions between investigated amino acid residues and terminal charges. To determine whether changing the protonation states of the N- and C-termini influence the conformational manifold of the central amino acid residue in tripeptides, we have examined the pH-dependence of unblocked trialanine and the conformational preferences of alanine in the alanine dipeptide. To this end, we measured and globally analyzed amide I' band profiles and NMR J-coupling constants. We described conformational distributions as the superposition of two-dimensional Gaussian distributions assignable to specific sub-spaces of the Ramachandran plot. Results show that the conformational ensemble of trialanine as a whole, and the pPII content (χpPII=0.84) in particular, remain practically unaffected by changing the protonation state. We found that compared to trialanine, the alanine dipeptide has slightly lower pPII content (χpPII=0.74) and an ensemble more reminiscent of the unblocked Gly-Ala-Gly model peptide. In addition, a two-state thermodynamic analysis of the conformational sensitive Δε(T) and 3J(HNHα)(T) data obtained from electronic circular dichroism and H-NMR spectra indicate that the free energy landscape of trialanine is similar in all protonation states. MD simulations for the investigated peptides corroborate this notion and show further that the hydration shell around unblocked trialanine is unaffected by the protonation/deprotonation of the C-terminal group. In contrast, the alanine dipeptide shows a reduced water density around the central residue as well as a less ordered hydration shell, which decreases the pPII propensity and reduces the lifetime of sampled conformations.The Journal of Physical Chemistry B 02/2013; · 3.70 Impact Factor