Bovine carbonic anhydrase (BCA) and its derivative with all lysine groups acetylated (BCA-Ac18) have different stabilities toward denaturation by sodium dodecyl sulfate (SDS). This difference is kinetic: BCA-Ac18 denatures more slowly than BCA by several orders of magnitude over concentrations of SDS ranging from 2.5 to 10 mM. The rates of renaturation of BCA-Ac18 are greater than those of BCA, when these proteins are allowed to refold from a denatured state ([SDS] = 10 mM) to a folded state ([SDS] = 0.1 to 1.5 mM). On renaturation, the yields of the correctly folded protein (either BCA or BCA-Ac18) decrease with increasing concentration of SDS. At intermediate concentrations of SDS (from 0.7 to 2 mM for BCA, and from 1.5 to 2 mM for BCA-Ac18), both unfolding and refolding of the proteins are too slow to be observed; an alternative processprobably aggregationcompetes with refolding of the denatured proteins at those intermediate concentrations. Because it is experimentally impractical to prove equilibrium, it is not possible to establish whether there is a difference in the thermodynamics of unfolding/refolding between BCA and BCA-Ac18.
Current-carrying microcircuits can generate strong magnetic-field gradients; these gradients, in turn, can control the position of magnetic microbeads in aqueous suspension. Micromagnetic systems were prepared using two representative soft lithography techniques—micromolding in capillaries and microtransfer molding—combined with electrodeposition. They can capture microbeads from solution, hold them in a fixed position, and move them along complex paths. Simply changing the current flowing in the systems can dynamically reconfigure the paths. The manipulation of magnetic microbeads using microfabricated circuits should expand current sample handling capabilities for biotechnology and combinational technology with or without the presence of net fluid flow and microfluidic channels.
[1] Material contrasts across faults are a common occurrence, and it is important to understand if these material contrasts can influence the path of rupture propagation. Here we examine models, solved numerically, of rupture propagation through one type of geometric complexity, that of a fault branch stemming from a planar main fault on which rupture initiates. This geometry, with a material contrast across the main fault, could be representative of either a mature strike-slip fault or a subduction zone interface. We consider branches in both the compressional and extensional quadrants of the fault, and material configurations in which the branch fault is in either the stiffer or the more compliant material as well as configurations with no material contrast. We find that there are regimes in which this elastic contrast can influence the rupture behavior at a branching junction, but there are also stress states for which the branch activation will not depend on the orientation of the mismatch. For the scenarios presented here, both compressional and extensional side branches are more likely to rupture if the branch is on the side of the fault with the more compliant material versus the stiffer material. The stresses induced on the branch fault, by rupture traveling on the main fault, are different for the two orientations of material contrast. We show how the interactions between rupture on the two faults determine which faults are activated.
As described in the previous chapter, the computation of energy release rates and stress intensity factors using the finite element method requires specification of various numerical parameters. These parameters relate to both the FE model itself (basically, the mesh) and the calculations performed during postprocessing of the FE results (basically, the calculation of the stiffness derivitive). In this chapter, the powerful automation of the LS-FEA framework is exploited to conduct extensive parametric studies, which illustrate the impact of these parameters on computational accuracy. The outcomes of these studies are used as the basis to establish recommended practices and default values for the associated numerical parameters, such as those in Table 16.1.
This paper describes the use of the OPTCOL (optically controlled collision) assay, an assay that uses optical tweezers to cause two biologically relevant particles to collide, to measure the potency of inhibitors that block the adhesion of wheat germ agglutinin (WGA) to the surface of erythrocytes. WGA was attached covalently to polystyrene microspheres (3 μm in diameter). Optical tweezers were used to cause an erythrocyte and a WGA-coated microsphere to collide in buffer at a controlled velocity. In the absence of inhibitor, or at low concentrations of soluble inhibitor, the microsphere adhered to the cell through polyvalent, biospecific interactions between WGA and N-acetylglucosamine (GlcNAc) and N-acetylneuraminic acid (NeuAc) groups present on the surface of erythrocyte. At high concentrations of soluble inhibitors, adhesion was inhibited. The potency of inhibition was quantified by measuring the probability of adhesion as a function of the concentration of the inhibitor. The inhibition constants derived from measurements using OPTCOL agreed well with those obtained in hemagglutination inhibition assays, and they were also close to the dissociation constants measured by isothermal titration calorimetry. The experimental data suggest that the binding of WGA to erythrocyte is cooperative, but that the binding of WGA to soluble ligand is not. The ability to examine dynamic adhesion in a highly controlled fashion makes OPTCOL a useful bioassay with which to study inhibition of protein-cell adhesion under biologically relevant conditions.
