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 NH3(+) 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.

Randomizing the Unfolded State of Peptides (and Proteins) by Nearest Neighbor Interactions between Unlike Residues

“…Previous studies by us4j,k,n,o, 7d, 13, 14 and others4c,g,l,m, 5a,b, 10b, 15 have clearly shown that, within the unfolded state, amino acid residues have unique and restricted conformational biases. Here we examine the mutual effect of unlike residues (x and y, Table 1) on residue‐level conformational bias through a combined analysis of vibrational and NMR results for GxyG peptide motifs.…”

“…A limited set of recent experiments on homotripeptides in our laboratories revealed that interactions between like‐residues in AAA, VVV, DDD and KKK lead to a further stabilization of the already preferred conformation of the central residue. For instance, the relatively high intrinsic β‐strand propensity found for valine is increased further in VVV,9 whereas the high pPII propensity for alanine and lysine is further increases in AAA5e and KKK 7d. In addition, the unusually high turn preference for protonated aspartic acid are further increased for the central residue in DDD relative to GDG 6a.…”

pH-Independence of Trialanine and the Effects of Termini Blocking in Short Peptides: A Combined Vibrational, NMR, UVCD, and Molecular Dynamics Study

“…5-7, 10, 24-26, 47-49, 89 Experimental work on e.g. AAA, the classical model system of unblocked tripeptides, essentially agrees in suggesting a large pPII content of its conformational distribution.…”

“…3-5, 10-12, 20-22 Moreover, it has become clear that amino acid residues show different conformational distributions, which can be altered by the nearest neighbors. 3, 22-26 In this context, polyproline II (pPII) has emerged as the dominant conformation for alanine, 10 whereas the pPII propensity of other residues is still a matter of a controversial debate. 3, 10, 11, 27 The canonical pPII conformation with (φ,ψ)=(−75°, 150°) is adopted by residues in trans -polyproline where it brings about a 3 1 -helix structure of the peptide.…”

Probing conformational propensities of histidine in different protonation states of the unblocked glycyl‐histidyl‐glycine peptide by vibrational and NMR spectroscopy

“…In the current study we combine IR, polarized Raman, VCD and 1 H NMR to examine the conformational preferences of histidine in different protonation states of GHG . Our data confirm that H has a comparatively high preference for β‐strand structures, which is only slightly modified by the protonation of the C‐terminal carboxylate group at room temperature.…”