243
8 Molecular Dynamics Simulations of Lipid Bilayers with Incorporated Peptides
We applied high-performance liquid chromatography (HPLC), CD, differential
scanning calorimetry DSC and attenuated total reflectance ATR-FTIR methods for
study studied specially designed α-helical transmembrane peptides (acetyl-K 2 -L m -
A n -K 2 -amide, where m + n = 24) in respect of their solution behavior and interactions
with phospholipids [19]. These peptides exhibit strong α-helical conformation in
water, membrane-mimetic media and lipid model membranes, however the stability
of the helices decreases as the Leu content decreases. Also, their binding to reversed
phase high-performance liquid chromatography columns is largely determined by
their hydrophobicity and the binding generally decreases with decrease in the Leu/
Ala ratio. However, the retention of these peptides by such columns is also affected
by the distribution of hydrophobic residues on their helical surfaces, being further
enhanced when peptide helical hydrophobic moments are increased by clustering
hydrophobic residues on one side of the helix. This clustering of hydrophobic residues also increases peptide propensity for self-aggregation in aqueous media and
enhances partitioning of the peptide into lipid bilayer membranes. We also found
that the peptides LA 3 LA 2 (acetyl-K 2 -(LA 3 LA 2 ) 3 LA 2 -K 2 -amide) and particularly
LA 6 (acetyl-K 2 -(LA 6 ) 3 LA 2 -K 2 -amide) associate less strongly with bilayer and perturb the thermotropic phase behavior of phosphatidylcholine bilayers much less
than peptides with higher L/A ratios. These results are consistent with free energies
calculated for the partitioning of these peptides between water and phospholipid bilayers. This suggests that LA 3 LA 2 has an equal tendency to partition into water and
into the hydrophobic core of phospholipid model membranes, whereas LA 6 should
strongly prefer the aqueous phase. We conclude that for α-helical peptides of this
type, Leu/Ala ratios of greater than 7/17 are required for stable transmembrane associations with phospholipid bilayers. Experimental studies have been focused also
on the analysis of the effect of substitution of some amino acids in peptides on their
properties. Idiong et al. [20] studied α-helical antimicrobial peptides purified from
the venom of the Central Asian spider Lachesana tarabaevi and showed that replacing the glycine at position 11 with alanine resulted in more rigid peptide structure
due to the reduced conformational flexibility.
Detailed DSC, FTIR, NMR and electron paramagnetic resonance (EPR) studies
of interaction of P 24 or L 24 with BLM [21] have revealed that the results obtained
from different physical techniques generally agree well with one another. However,
certain discrepancies have been found in comparison of the results obtained by spectroscopic techniques, i.e. FTIR and
2
H-NMR. While the
2
H-NMR technique indicated that incorporation of P 24 peptide into the DPPC bilayers resulted in a decrease
of the ordering of the membrane in gel state and increase in the liquid crystalline
(LC) state, FTIR experiments suggest that peptide induced a decrease of the ordering of the lipid bilayer in both structural state of the membrane [21]. This discrepancy has been explained by different peculiarities of these two methods. While the
order parameters in
2
H-NMR spectroscopy are primarily sensitive to trans/gauche
isomerisation, the molecular interpretation of the changes in membrane ordering
based on changes in frequency of the methylene stretching modes in IR spectroscopy are likely attributed to the sensitivity of the band position phenomena other than
trans/gauche isomerisation, such as the interchain coupling and the contribution
8 Molecular Dynamics Simulations of Lipid Bilayers with Incorporated Peptides
We applied high-performance liquid chromatography (HPLC), CD, differential
scanning calorimetry DSC and attenuated total reflectance ATR-FTIR methods for
study studied specially designed α-helical transmembrane peptides (acetyl-K 2 -L m -
A n -K 2 -amide, where m + n = 24) in respect of their solution behavior and interactions
with phospholipids [19]. These peptides exhibit strong α-helical conformation in
water, membrane-mimetic media and lipid model membranes, however the stability
of the helices decreases as the Leu content decreases. Also, their binding to reversed
phase high-performance liquid chromatography columns is largely determined by
their hydrophobicity and the binding generally decreases with decrease in the Leu/
Ala ratio. However, the retention of these peptides by such columns is also affected
by the distribution of hydrophobic residues on their helical surfaces, being further
enhanced when peptide helical hydrophobic moments are increased by clustering
hydrophobic residues on one side of the helix. This clustering of hydrophobic residues also increases peptide propensity for self-aggregation in aqueous media and
enhances partitioning of the peptide into lipid bilayer membranes. We also found
that the peptides LA 3 LA 2 (acetyl-K 2 -(LA 3 LA 2 ) 3 LA 2 -K 2 -amide) and particularly
LA 6 (acetyl-K 2 -(LA 6 ) 3 LA 2 -K 2 -amide) associate less strongly with bilayer and perturb the thermotropic phase behavior of phosphatidylcholine bilayers much less
than peptides with higher L/A ratios. These results are consistent with free energies
calculated for the partitioning of these peptides between water and phospholipid bilayers. This suggests that LA 3 LA 2 has an equal tendency to partition into water and
into the hydrophobic core of phospholipid model membranes, whereas LA 6 should
strongly prefer the aqueous phase. We conclude that for α-helical peptides of this
type, Leu/Ala ratios of greater than 7/17 are required for stable transmembrane associations with phospholipid bilayers. Experimental studies have been focused also
on the analysis of the effect of substitution of some amino acids in peptides on their
properties. Idiong et al. [20] studied α-helical antimicrobial peptides purified from
the venom of the Central Asian spider Lachesana tarabaevi and showed that replacing the glycine at position 11 with alanine resulted in more rigid peptide structure
due to the reduced conformational flexibility.
Detailed DSC, FTIR, NMR and electron paramagnetic resonance (EPR) studies
of interaction of P 24 or L 24 with BLM [21] have revealed that the results obtained
from different physical techniques generally agree well with one another. However,
certain discrepancies have been found in comparison of the results obtained by spectroscopic techniques, i.e. FTIR and
2
H-NMR. While the
2
H-NMR technique indicated that incorporation of P 24 peptide into the DPPC bilayers resulted in a decrease
of the ordering of the membrane in gel state and increase in the liquid crystalline
(LC) state, FTIR experiments suggest that peptide induced a decrease of the ordering of the lipid bilayer in both structural state of the membrane [21]. This discrepancy has been explained by different peculiarities of these two methods. While the
order parameters in
2
H-NMR spectroscopy are primarily sensitive to trans/gauche
isomerisation, the molecular interpretation of the changes in membrane ordering
based on changes in frequency of the methylene stretching modes in IR spectroscopy are likely attributed to the sensitivity of the band position phenomena other than
trans/gauche isomerisation, such as the interchain coupling and the contribution