We report a point mutation in the second contact shell of the high-affinity streptavidin-biotin complex that appears to reduce binding affinity through transmitted effects on equilibrium dynamics. The Y54F streptavidin mutation causes a 75-fold loss of binding affinity with 73-fold faster dissociation, a large loss of binding enthalpy (ΔΔH, 3.4 kcal/mol at 37 °C) and a small gain in binding entropy (TΔΔS, 0.7 kcal/mol). The removed Y54 hydroxyl is replaced by a water molecule in the bound structure, but there are no observable changes in structure in the first contact shell and no additional changes surrounding the mutation. Molecular dynamics simulations reveal a large increase in atomic fluctuations for W79, a key biotin contact residue, compared to the wild type complex. The increased W79 fluctuations are caused by loss of water-mediated hydrogen bonds between the Y54 hydroxyl group and peptide backbone atoms in and near W79. We propose that the increased fluctuations diminish the integrity of the W79-biotin interaction and represent a loosening of the “tryptophan collar” which is critical to the slow dissociation and high affinity of streptavidin-biotin binding. These results illustrate how changes in protein dynamics distal to the ligand binding pocket can have a profound impact on ligand binding, even when equilibrium structure is unperturbed.

We prepare a temperature-dependent formulation of the integrated kinetics for the overall process of photosynthesis in eukaryotic cells. To avoid complexity, the C4 plants are chosen because their rate of photosynthesis is independent of the partial pressure of O2. A systematically simplified but comprehensive scheme for both light and dark reactions is considered. The reaction rate per reaction center in the thylakoid membrane is related to the rate of exciton transfer between chlorophyll neighbors. An expression is formulated for the light reaction rate (R1'). The NADPH formation rate is related to R1' and the survival probability of the membrane. Rates of different steps in the simplified scheme can be related to each other by applying a few steady state conditions. The saturation probability of CO2 in a bundle sheath is also considered. The photochemical efficiency (phi) appears in terms of these probabilities. We find the glucose production rate as R(glucose) = (8/3) upsilon L: [corrected] R1'phi g(T)([G3P]/[P(i)]2) exp(-deltaG(E)S/RT), where g(T) is the activation quotient of the involved enzymes, G3P and P(i) represent glyceraldehyde-3-phosphate and inorganic phosphates, and deltaG(E)S is the free energy for the apparent equilibrium between G3P and glucose. This is the first time that such a comprehensive expression for R(glucose) has been derived. The probabilities are generally given by sigmoid curves. The corresponding parameters can be easily determined. The quotient g(T) incorporates a Gaussian distribution for temperature dependence and a sigmoid function describing deactivation. The theoretical plots of photochemical efficiency and glucose production rate versus temperature are in excellent agreement with the experimental ones, thereby validating the formalism.

A physicochemical interpretation of a recently formulated temperature-dependent, steady-state rate expression for the production of glucose equivalent in C(4) plants is given here. We show that the rate equation is applicable to a wide range of C(4) plants.

No abstract

We give a detailed description of the use of explicit as well as implicit solvation treatments to compute the reduction potentials of biomolecules in a medium. The explicit solvent method involves quantum mechanical/molecular mechanics (QM/MM) treatment of the solvated moiety followed by a Monte-Carlo (MC) simulation of the primary solvent layer. The QM task for considerably large biomolecules is normally carried out by density functional treatment (DFT) along with the MM-assisted evaluation of the most stable configuration for the primary layer and biomolecule complex. The MC simulation accounts for the dynamics of the associated solvent molecules. Contributions of the solvent molecules of the bulk towards the absolute free energy change of the reductive process are incorporated in terms of the Born energy of ion-dielectric interaction, the Onsager energy of dipole-dielectric interaction and the Debye-Hückel energy of ion-ionic cloud interaction. In the implicit solvent treatment, one employs the polarizable continuum model (PCM). Thus the contribution of all the solvent molecules towards the free energy change are incorporated by considering the whole solvent as a dielectric continuum.As an example, the QM(DFT)/MM/MC-Born/Onsager/Debye-Hückel corrections yielded the oneelectron reduction potential of Pheophytin-a in the solvent DMF as −0⋅92 ± 0⋅27 V and the two-electron reduction potential as −1⋅34 ± 0⋅25 V at 298⋅15 K while the DFT-DPCM method yielded the corresponding values as −1⋅03 ± 0⋅17 V and −1⋅30 ± 0⋅17 V, respectively. The calculated values more or less agree with the observed mid-point potentials of −0⋅90 V and −1⋅25 V, respectively. Moreover, a numerical finite difference Poisson-Boltzmann solution along with the DFT-DPCM methodology was employed to calculate the reduction potential of Pheophytin-a within the thylakoid membrane. The calculated reduction potential value of −0⋅58 V is in agreement with the reported value of −0⋅61 V that appears in the socalled Z-scheme and is considerably different from the value in vitro.