Relative stability of the open and closed conformations of the active site loop of streptavidin

The eight-residue surface loop, 45-52 (Ser, Ala, Val, Gly, Asn, Ala, Glu, Ser), of the homotetrameric protein streptavidin has a "closed" conformation in the streptavidin-biotin complex, where the corresponding binding affinity is one of the strongest found in nature (ΔG ～ -18 kcal∕mol). However, in most of the crystal structures of apo (unbound) streptavidin, the loop conformation is "open" and typically exhibits partial disorder and high B-factors. Thus, it is plausible to assume that the loop structure is changed from open to closed upon binding of biotin, and the corresponding difference in free energy, ΔF = F(open) - F(closed) in the unbound protein, should therefore be considered in the total absolute free energy of binding. ΔF (which has generally been neglected) is calculated here using our "hypothetical scanning molecular-dynamics" (HSMD) method. We use a protein model in which only the atoms closest to the loop are considered (the "template") and they are fixed in the x-ray coordinates of the free protein; the x-ray conformation of the closed loop is attached to the same (unbound) template and both systems are capped with the same sphere of TIP3P water. Using the force field of the assisted model building with energy refinement (AMBER), we carry out two separate MD simulations (at temperature T = 300 K), starting from the open and closed conformations, where only the atoms of the loop and water are allowed to move (the template-water and template-loop interactions are considered). The absolute F(open) and F(closed) (of loop + water) are calculated from these trajectories, where the loop and water contributions are obtained by HSMD and a thermodynamic integration (TI) process, respectively. The combined HSMD-TI procedure leads to total (loop + water) ΔF = -27.1 ± 2.0 kcal∕mol, where the entropy TΔS constitutes 34% of ΔF, meaning that the effect of S is significant and should not be ignored. Also, ΔS is positive, in accord with the high flexibility of the open loop observed in crystal structures, while the energy ΔE is unexpectedly negative, thus also adding to the stability of the open loop. The loop and the 250 capped water molecules are the largest system studied thus far, which constitutes a test for the efficiency of HSMD-TI; this efficiency and technical issues related to the implementation of the method are also discussed. Finally, the result for ΔF is a prediction that will be considered in the calculation of the absolute free energy of binding of biotin to streptavidin, which constitutes our next project.     

Secondary electron emission control in X-ray photoelectron spectroscopy

Secondary electron emission (SEE) is a major player in surface charging during X-ray photoelectron spectroscopy (XPS); its characteristics and applicability as a current source for electrical measurements are studied. We employ sample biasing and a top retarding grid to control the photoelectron current, and further compare their I–V characteristics with direct spectroscopy of the secondary electrons. Using silica-coated gold substrates, the effect of sample work function on the emitted secondary electrons is shown and fine control over the surface potential gradients, in the range of 10–100 meV, is achieved. XPS-based chemically resolved electrical measurements (CREM) can thus be extended to the positive current regime.     

A general substructure synthesis method for the dynamic simulation of complex structures

In this paper a general substructure synthesis method is developed for the dynamic analysis of complex flexible structures. The motion of each substructure is represented by a given number of substructure admissible functions. Substructure admissible functions are often low-order polynomials, and hence computationally easy to work with. The otherwise disjoint substructures are connected together to form a whole structure by imposing approximate geometric compatibility conditions by means of the method of weighted residuals. The behavior of the estimated eigenvalues obtained by the substructures synthesis method can be ascertained by means of a bracketing theorem. The estimated eigenvalues do converge to the actual eigenvalues of the original structure, although it is necessary to consider two limiting processes, one in which the number of substructure admissible functions is increased and the other in which the number of internal boundary weighting functions is increased.     

Energy level and band alignment for GaAs-alkylthiol monolayer-Hg junctions from electrical transport and photoemission experiments

A series of p- and n-GaAs-S-C(n)H(2n+1) || Hg junctions are prepared, and the electronic transport through them is measured. From current-voltage measurements, we find that, for n-GaAs, transport occurs by both thermionic emission and tunneling, with the former dominating at low forward bias and the latter dominating at higher forward bias. For p-GaAs, tunneling dominates at all bias voltages. By combining the analysis of the transport data with results from direct and inverse photoemission spectroscopy, we deduce an energy band diagram of the system, including the tunnel barrier and, with this barrier and within the Simmons tunneling model, extract an effective mass value of 1.5-1.6m(e) for the electronic carriers that cross the junctions. We find that transport is well-described by lowest unoccupied and highest occupied states at 1.3-1.4 eV above and 2.0-2.2 eV below the Fermi level. At the same time, the photoemission data indicate that there are continua of states from the conduction band minimum and the valence band maximum, the density of which varies with energy. On the basis of our results, it appears likely that, for both types of junctions, electrons are the main carrier type, although holes may contribute significantly to the transport in the p-GaAs system.     

Minimalist explicit solvation models for surface loops in proteins

We have performed molecular dynamics simulations of protein surface loops solvated by explicit water, where a prime focus of the study is the small numbers (e.g., ~100) of explicit water molecules employed. The models include only part of the protein (typically 500 - 1000 atoms), and the water molecules are restricted to a region surrounding the loop. In this study, the number of water molecules (N(w)) is systematically varied, and convergence with large N(w) is monitored to reveal N(w)(min), the minimum number required for the loop to exhibit realistic (fully hydrated) behavior. We have also studied protein surface coverage, as well as diffusion and residence times for water molecules as a function of N(w). A number of other modeling parameters are also tested. These include the number of environmental protein atoms explicitly considered in the model, as well as two ways to constrain the water molecules to the vicinity of the loop (where we find one of these methods to perform better when N(w) is small). The results (for RMSD and its fluctuations for four loops) are further compared to much larger, fully solvated systems (using ~10,000 water molecules under periodic boundary conditions and Ewald electrostatics), and to results for the GBSA implicit solvation model. We find that the loop backbone can stabilize with a surprisingly small number of water molecules (as low as 5 molecules per amino acid residue). The side chains of the loop require somewhat larger N(w), where the atomic fluctuations become too small if N(w) is further reduced. Thus, in general, we find adequate hydration to occur at roughly 12 water molecules per residue. This is an important result, because at this hydration level, computational times are comparable to those required for GBSA. Therefore these "minimalist explicit models" can provide a viable and potentially more accurate alternative. The importance of protein loop modeling is discussed in the context of these, and other, loop models, along with other challenges including the relevance of appropriate free energy simulation methodology for assessment of conformational stability.     

Chemically resolved photovoltage measurements in CdSe nanoparticle films

Light-induced chemically resolved electrical measurements (CREM) under controlled electrical conditions are used to study photovoltaic effects at selected regions in nanocrystalline CdSe-based films. The method, based on X-ray photoelectron spectroscopy (XPS), possesses unique capabilities for exploring charge trapping and charge transport mechanisms, combining spectrally filtered input signals with photocurrent detection and with a powerful, site-selective, photovoltage probe.     

Multicanonical procedure for continuum peptide models

The multicanonical (Muca) Monte Carlo method enables simulating a system over a wide range of temperatures and thus has become an efficient tool for studying spin glasses, first‐order phase transitions, the helix–coil transition of polypeptides, and protein folding. However, implementation of the method requires calculating the multicanonical weights by an iterative procedure that is not straightforward and is a stumbling block for newcomers. A recursive procedure that takes into account the statistical errors of all previous iterations and thus enables an automatic calculation of the weights without the need for human intervention after each iteration has been proposed. This procedure, which has already been tested successfully for lattice systems, is extended here to continuum models of peptides and proteins. The method is examined in detail and tested for models of the pentapeptide Leu‐enkephalin (Tyr‐Gly‐Gly‐Phe‐Leu) described by the potential energy function ECEPP. Because of the great interest in the structural mapping of the low‐energy region of biomolecules, the energy of structures selected from the Muca trajectory is minimized. The extent of conformational coverage provided by the method is examined and found to be very satisfactory. © 2000 John Wiley & Sons, Inc. J Comput Chem 21: 1251–1261, 2000     

The role played by oxygen plasma on Teflon: relevance to the concept of "cryptoelectrons"

Dr Williams (AIP Adv., 2012, 2, 010701) suggested that cleaning Teflon by high pressure oxygen plasma may have affected our result that Cu(2+) and Pd(2+) ions can be absorbed but not chemically reduced by a Teflon surface rubbed by PMMA (Phys. Chem. Chem. Phys., 2012, 14, 5551). In response, we show that this treatment does not affect the adsorption of Cu(2+) and Pd(2+). We reaffirm our statement that Cu(2+) and Pd(2+) ions can be adsorbed by a Teflon surface only after rubbing with the organic polymers, not before rubbing.