Molecular Modeling and Affinity Determination of scFv Antibody: Proper Linker Peptide Enhances Its Activity

One of existing strategies to engineer active antibody is to link VH and VL domains via a linker peptide. How the composition, length, and conformation of the linker affect antibody activity, however, remains poorly understood. In this study, a dual approach that coordinates molecule modeling, biological measurements, and affinity evaluation was developed to quantify the binding activity of a novel stable miniaturized anti-CD20 antibody or singlechain fragment variable (scFv) with a linker peptide. Upon computer-guided homology modeling, distance geometry analysis, and molecular superimposition and optimization, three new linker peptides PT1, PT2, and PT3 with respective 7, 10, and 15 residues were proposed and three engineered antibodies were then constructed by linking the cloned VH and VL domains and fusing to a derivative of human IgG1. The binding stability and activity of scFv-Fc chimera to CD20 antigen was quantified using a micropipette adhesion frequency assay and a Scatchard analysis. Our data indicated that the binding affinity was similar for the chimera with PT2 or PT3 and ~24-fold higher than that for the chimera with PT1, supporting theoretical predictions in molecular modeling. These results further the understanding in the impact of linker peptide on antibody structure and activity.

Topics in vortex motion

Six topics in incompressible, inviscid fluid flow involving vortex motion are presented. The stability of the unsteady flow field due to the vortex filament expanding under the influence of an axial compression is examined in the first chapter as a possible model of the vortex bursting observed in aircraft contrails. The filament with a stagnant core is found to be unstable to axisymmetric disturbances. For initial disturbances with the form of axisymmetric Kelvin waves, the filament with a uniformly rotating core is neutrally stable, but the compression causes the disturbance to undergo a rapid increase in amplitude. The time at which the increase occurs is, however, later than the observed bursting times, indicating the bursting phenomenon is not caused by this type of instability.

In the second and third chapters the stability of a steady vortex filament deformed by two-dimensional strain and shear flows, respectively, is examined. The steady deformations are in the plane of the vortex cross-section. Disturbances which deform the filament centerline into a wave which does not propagate along the filament are shown to be unstable and a method is described to calculate the wave number and corresponding growth rate of the amplified waves for a general distribution of vorticity in the vortex core.

In Chapter Four exact solutions are constructed for two-dimensional potential flow over a wing with a free ideal vortex standing over the wing. The loci of positions of the free vortex are found and the lift is calculated. It is found that the lift on the wing can be significantly increased by the free vortex.

The two-dimensional trajectories of an ideal vortex pair near an orifice are calculated in Chapter Five. Three geometries are examined, and the criteria for the vortices to travel away from the orifice are determined.

Finally, Chapter Six reproduces completely the paper, "Structure of a linear array of hollow vortices of finite cross-section," co-authored with G. R. Baker and P. G. Saffman. Free streamline theory is employed to construct an exact steady solution for a linear array of hollow, or stagnant cored vortices. If each vortex has area A and the separation is L, then there are two possible shapes if A^(1/2)/L is less than 0.38 and none if it is larger. The stability of the shapes to two-dimensional, periodic and symmetric disturbances is considered for hollow vortices. The more deformed of the two possible shapes is found to be unstable, while the less deformed shape is stable.