Lee Brent L, Kuczera Krzysztof, Middaugh C Russell, Jas Gouri S
Department of Chemistry, The University of Kansas, Lawrence, Kansas 66045, USA.
Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, Kansas 66047, USA.
J Chem Phys. 2016 Jun 28;144(24):245103. doi: 10.1063/1.4954241.
The time-resolved parallel artificial membrane permeability assay with fluorescence detection and comprehensive computer simulations are used to study the passive permeation of three aromatic dipeptides-N-acetyl-phenylalanineamide (NAFA), N-acetyltyrosineamide (NAYA), and N-acetyl-tryptophanamide (NATA) through a 1,2-dioleoyl-sn-glycero-3-phospocholine (DOPC) lipid bilayer. Measured permeation times and permeability coefficients show fastest translocation for NAFA, slowest for NAYA, and intermediate for NATA under physiological temperature and pH. Computationally, we perform umbrella sampling simulations to model the structure, dynamics, and interactions of the peptides as a function of z, the distance from lipid bilayer. The calculated profiles of the potential of mean force show two strong effects-preferential binding of each of the three peptides to the lipid interface and large free energy barriers in the membrane center. We use several approaches to calculate the position-dependent translational diffusion coefficients D(z), including one based on numerical solution the Smoluchowski equation. Surprisingly, computed D(z) values change very little with reaction coordinate and are also quite similar for the three peptides studied. In contrast, calculated values of sidechain rotational correlation times τrot(z) show extremely large changes with peptide membrane insertion-values become 100 times larger in the headgroup region and 10 times larger at interface and in membrane center, relative to solution. The peptides' conformational freedom becomes systematically more restricted as they enter the membrane, sampling α and β and C7eq basins in solution, α and C7eq at the interface, and C7eq only in the center. Residual waters of solvation remain around the peptides even in the membrane center. Overall, our study provides an improved microscopic understanding of passive peptide permeation through membranes, especially on the sensitivity of rotational diffusion to position relative to the bilayer.
采用具有荧光检测功能的时间分辨平行人工膜通透性测定法以及全面的计算机模拟，研究三种芳香族二肽——N-乙酰苯丙氨酸酰胺（NAFA）、N-乙酰酪氨酸酰胺（NAYA）和N-乙酰色氨酸酰胺（NATA）通过1,2-二油酰基-sn-甘油-3-磷酸胆碱（DOPC）脂质双层的被动渗透。在生理温度和pH条件下，测得的渗透时间和渗透系数表明，NAFA的转运速度最快，NAYA最慢，NATA居中。在计算方面，我们进行伞形采样模拟，以模拟肽的结构、动力学及其与脂质双层距离z的函数关系。计算得到的平均力势分布图显示出两种强烈的效应——三种肽中的每一种都优先结合到脂质界面，以及在膜中心存在较大的自由能垒。我们使用几种方法来计算位置依赖性平移扩散系数D(z)，其中一种基于对Smoluchowski方程的数值求解。令人惊讶的是，计算得到的D(z)值随反应坐标变化很小，并且对于所研究的三种肽也非常相似。相比之下，计算得到的侧链旋转相关时间τrot(z)值随肽插入膜的过程变化极大——相对于溶液，在头基区域的值增大100倍，在界面和膜中心增大10倍。随着肽进入膜内，其构象自由度系统性地受到更多限制，在溶液中采样α和β以及C7eq构象，在界面处采样α和C7eq构象，在膜中心仅采样C7eq构象。即使在膜中心，肽周围仍存在溶剂化残留水。总体而言，我们的研究对肽通过膜的被动渗透提供了更深入的微观理解，特别是对旋转扩散相对于双层位置的敏感性。
