1. Lactam bridge stabilization of alpha-helical peptides: ring size, orientation and positional effects
M E Houston Jr, C L Gannon, C M Kay, R S Hodges J Pept Sci. 1995 Jul-Aug;1(4):274-82. doi: 10.1002/psc.310010408.
A series of 14 residue amphipathic alpha-helical peptides, in which the sidechains of glutamic acid and lysine have been covalently joined, was synthesized in order to determine the effect of spacing, position and orientation of these lactam bridges. It was found that although an (i, i+3) spacing would position the lactam bridge on the same face of the helix, these lactams with 18-member rings were actually helix-destabilizing regardless of position or location. On the other hand, (i, i+4) lactams with 21-member rings were helix-stabilizing but this was dependent on orientation. Glutamic acid-lysine lactams increased the helical content of the peptide when compared with their linear homologue in benign conditions (50 mM KH2PO4, 100 mM KCl, pH 7). Two Glu-Lys (i, i+4) lactams located at the N- and C-termini gave rise to a peptide with greater than 99% helical content in benign conditions. Peptides with Lys-Glu oriented lactams were random structures in benign conditions but in the presence of 50% TFE could be induced into a helical conformation. The stability of the single-stranded alpha-helices, as measured by thermal denaturations in 25% TFE indicated that Glu-Lys oriented lactam bridges stabilized the helical conformation relative to the linear unbridged peptide. One Glu-Lys lactam in the middle of the peptide was more effective at stabilizing helical structure than two Glu-Lys lactams positioned one at each end of the molecule. The lactams with the Lys-Glu orientation were destabilizing relative to the unbridged peptide. This study demonstrates that correct orientation and position of a lactam bridge is critical in order to design peptides with high helical content in aqueous media.
2. [Lysine residue as an amino acid substitute for conformational constraint of Xaa-Asp fragment of biologically active peptides using side chain lactamization]
P V Kostetskiĭ, I V Artem'ev Bioorg Khim. 1997 Mar;23(3):168-73.
The model cyclopeptide Ac-Lys-Asp-NHMe was used to test Lys as a possible substitute for Xaa in peptide fragment Xaa-Asp whose conformational mobility would be constrained by lactamization of the Lys and Asp side chains. By means of theoretical conformational analysis, such a lactam was shown to be capable of fixing several conformations of the peptide. Among them, 32 conformations corresponded to 8 low-energy regions of the linear peptide Ac-Ala-Asp-NHMe, which was chosen as a model for the peptide fragment Xaa-Asp. In this case, the conformational possibilities of the Xaa residue were constrained to two regions of the Phi, Psi-map, (A + G) and C according to Zimmermann-Sheraga notation.
3. Effect of lysine side chain length on intra-helical glutamate--lysine ion pairing interactions
Richard P Cheng, Prashant Girinath, Raheel Ahmad Biochemistry. 2007 Sep 18;46(37):10528-37. doi: 10.1021/bi700701z. Epub 2007 Aug 25.
Ion-pairing interactions are important for protein stabilization. Despite the apparent electrostatic nature of these interactions, natural positively charged amino acids Lys and Arg have multiple methylenes linking the charged functionality to the backbone. Interestingly, the amino acids Lys and Orn have positively charged side chains that differ by only one methylene. However, only Lys is encoded and incorporated into proteins. To investigate the effect of side chain length of Lys on ion-pairing interactions, a series of 12 monomeric alpha-helical peptides containing potential Glu-Xaa (i, i+3), (i, i+4) and (i, i+5) (Xaa = Lys, Orn, Dab, Dap) interactions were studied by circular dichroism (CD) spectroscopy at pH 7 and 2. At pH 7, no Glu-Xaa (i, i+5) interaction was observed, regardless of the Xaa side chain length. Furthermore, only Lys was capable of supporting Glu-Xaa (i, i+3) interactions, whereas any Xaa side chain length supported Glu-Xaa (i, i+4) interactions. Side chain conformational analysis by molecular mechanics calculations showed that the side chain length of Lys enables the Glu-Xaa (i, i+3) interaction with lower energy conformations compared to residues with side chain lengths shorter than that of Lys. Furthermore, these calculated low energy conformers were consistent with conformations of intra-helical Glu-Lys salt bridges in a non-redundant protein structure database. Importantly, the CD spectra for peptides with Glu-Lys interactions did not alter significantly upon changing the pH because of a greater contribution to these interactions by forces other than electrostatics. Incorporating side chains just one methylene shorter (Orn) resulted in significant pH dependence or lack of interaction, suggesting that nature has chosen Lys to form durable interactions with negatively charged functional groups.
