SMD Simulations Research Articles

Single Molecule Force Spectroscopy and Steered Molecular Dynamics Simulations Reveal the Mechanical Design of the Third FnIII Domain of Tenascin-C

By combining single molecule atomic force microscopy, proline mutagenesis and steered molecular dynamics simulations, we investigate the mechanical unfolding dynamics and mechanical design of the third FnIII domain of tenascin-C (TNfn3) in detail. The mechanical stability of TNfn3 is found to be similar to that of other constituting FnIII domains of tenascin-C, and the unfolding process of TNfn3 is an apparent two-state process. The hydrophobic core packing of TNfn3 was previously reported as the key element of its mechanical stability. Here, employing proline mutagenesis to block the formation of backbone hydrogen bonds and introduce structural disruption in β sheet, we showed that not only hydrophobic core packing plays important roles in determining the mechanical stability of TNfn3, backbone hydrogen bonds in β hairpins are also responsible for the overall mechanical stability of TNfn3. Furthermore, proline mutagenesis revealed that the mechanical design of TNfn3 is very robust and proline substitution in β sheets only leads to mild reduction in mechanical stability. We also compare the AFM results with those of SMD simulations to understand the molecular details underlying the mechanical unfolding of TNfn3. We found that the mechanical unfolding and design of TNfn3 is significantly different from its structural homologue the tenth FnIII domain from fibronectin. These results serve as a starting point for systematically analyzing the mechanical architecture of other FnIII domains in tenascins-C and will help to gain a better understanding of some of the complex features observed for the stretching of native tenascin-C.

Behavior Regulation of Adsorbed Proteins via Hydroxyapatite Surface Texture Control

The influence of nanometer-scale interfaces on proteins has received much attention in recent years. The dynamic behaviors of bone morphogenetic protein-7 (BMP-7) on a series of hydroxyapatite (HAP) surface textures were investigated to explore the influence of different surface textures using molecular dynamics (MD), steered molecular dynamics simulations (SMD), and quantum mechanics calculations. It is observed that the interaction energy curve from SMD simulations can exhibit the dynamic behavior of BMP-7 in detail. Both the type and the number difference of the adsorptive residues and the intensity discrepancy of interaction, which is induced by the specific texture of the HAP surface, could be uncovered from the energy curve qualitatively and semiquantitatively in this study. The largest conformational change occurs in the system 010+a. The quantum mechanics calculations suggest that there is a phenomenon of electron transfer from HAP to the groups of BMP-7 during the adsorption process. These findings suggest that surface-engineering techniques could be employed to directly control the texture of HAP surfaces in order to regulate the behavior of a protein adsorbed onto the nanometer-scale interface.

Direct Detection of the Formation of V-Amylose Helix by Single Molecule Force Spectroscopy

An important polysaccharide, amylose crystallizes as a regular single left-handed helix from a propanol, butanol, or iodine solution. However, its solution structure remains elusive because amylose does not form molecular solutions in these solvents, and standard spectroscopic techniques cannot be exploited to determine its structure. Using AFM, we forced individual amylose chains adsorbed to a surface to enter these poor solvents and carried out stretch-release measurements on them in solution. In this manner, we directly captured the formation of individual amylose helices induced by butanol and iodine. With an accuracy approaching that of X-ray diffraction on amylose crystals, we determined that the pitch of the helix in solution is 1.3 angstroms/ring. We also directly measured the force driving the formation of the helix in solution to be 50 pN. SMD simulations in explicit butanol reproduced the AFM-measured force-extension curves and revealed that the long plateau feature is caused by the rupture of O(2)n-O(6)(n+6) and O(3)n-O(6)(n+6) hydrogen bonds and by the unwinding of the helix. We also found that amylose helices formed in iodine solution are more compliant and hysteretic as compared to helices in butanol, which extend/relax reversibly. In iodine solution, the formation of the helix is inhibited by force and limited by the slow kinetics of the amylose-iodine complex. By forcing individual molecules into poor solvents and performing force spectroscopy measurements in solution, our AFM approach uniquely supplements X-ray diffraction and NMR methods for investigating solution conformations of insoluble biopolymers.