Computer Simulation of Assembly and Co-operativity of Hexameric AAA ATPases

AAA ATPases form a functionally diverse superfamily of proteins. Most members form homo-hexameric ring complexes, are catalytically active only in the fully assembled state, and show co-operativity among the six subunits. The mutual dependence among the subunits is clearly evidenced by the fact that incorporation of mutated, inactive subunits can decrease the activity of the remaining wild type subunits. For the first time, we develop here models to describe this form of allostery, evaluate them in a simulation study, and test them on experimental data. We show that it is important to consider the assembly reactions in the kinetic model, and to define a formal inhibition scheme. We simulate three inhibition scenarios explicitly, and demonstrate that they result in differing outcomes. Finally, we deduce fitting formulas, and test them on real and simulated data. A non-competitive inhibition formula fitted experimental and simulated data best. To our knowledge, our study is the first one that derives and tests formal allosteric schemes to explain the inhibitory effects of mutant subunits on oligomeric enzymes.     

Structural consequences of mutation.

The way a protein responds to mutation provides key insights into its architecture and energetics. Mutations are improving the understanding both of protein folding and stability, and of the adaptability of the hydrophobic core. The importance of intermolecular effects in crystal structures is being emphasized and new insights into the correspondence between crystal and solution structures are being developed.     

An implantable transducer for measuring tension in an anterior cruciate ligament graft.

The goal of this study was to develop a new implantable transducer for measuring anterior cruciate ligament (ACL) graft tension postoperatively in patients who have undergone ACL reconstructive surgery. A unique approach was taken of integrating the transducer into a femoral fixation device. To devise a practical in vivo calibration protocol for the fixation device transducer (FDT), several hypotheses were investigated: (1) The use of a cable versus the actual graft as the means for applying load to the FDT during calibration has no significant effect on the accuracy of the FDT tension measurements; (2) the number of flexion angles at which the device is calibrated has no significant effect on the accuracy of the FDT measurements; (3) the friction between the graft and femoral tunnel has no significant effect on measurement accuracy. To provide data for testing these hypotheses, the FDT was first calibrated with both a cable and a graft over the full range of flexion. Then graft tension was measured simultaneously with both the FDT on the femoral side and load cells, which were connected to the graft on the tibial side, as five cadaver knees were loaded externally. Measurements were made with both standard and overdrilled tunnels. The error in the FDT tension measurements was the difference between the graft tension measured by the FDT and the load cells. Results of the statistical analyses showed that neither the means of applying the calibration load, the number of flexion angles used for calibration, nor the tunnel size had a significant effect on the accuracy of the FDT. Thus a cable may be used instead of the graft to transmit loads to the FDT during calibration, thus simplifying the procedure. Accurate calibration requires data from just three flexion angles of 0, 45, and 90 deg and a curve fit to obtain a calibration curve over a continuous range of flexion within the limits of this angle group. Since friction did not adversely affect the measurement accuracy of the FDT, the femoral tunnel can be drilled to match the diameter of the graft and does not need to be overdrilled. Following these procedures, the error in measuring graft tension with the FDT averages less than 10 percent relative to a full-scale load of 257 N.     

Pendelluft flow in symmetric airway bifurcations.

We propose a mathematical model for pendelluft flow in a single airway bifurcation. The model is motivated by an apparatus used in an experimental study of the pendelluft by Ultman et al. (1988). We derive differential equations governing the fluid flow, which directly connect physiological parameters to the variables determining the pendelluft; this approach allows us to include nonlinearity in the model. If nonlinearity is neglected, our model is identical to the R-I-C circuits used by previous investigators. If nonlinearity is retained, we show that pendelluft can occur even in perfectly symmetric airway bifurcations. For the specific apparatus used in the experiments of High et al. (1991), we demonstrate that two qualitatively different pendelluft flows can occur in the system.