Affinity recovery of eight HER2-binding affibody variants using an anti-idiotypic affibody molecule as capture ligand

Affibody molecules generated by combinatorial protein engineering to bind the human epidermal growth factor receptor 2 (HER2) have in earlier studies proven to be promising tracers for HER2-mediated molecular imaging of cancer. Amino acid extensions either at the N- or C-terminus of these Z(HER2) affibody molecules, have been successfully employed for site-specific radiolabeling of the tracer candidates. Hexahistidyls or other tags, which would be convenient for recovery purposes, should be avoided since they could negatively influence the tumor targeting efficacy and biodistribution properties of the tracer. Using a new ?-lactamase-based protein fragment complementation assay (PCA), an affibody molecule was isolated which bound a Z(HER2) affibody molecule with sub-micromolar affinity, but not unrelated affibody molecules. This suggests that the interacting area include the HER2-binding surface of Z(HER2). This novel anti-idiotypic affibody molecule Z(E01) was produced in Escherichia coli, purified, and chemically coupled to a chromatography resin in order to generate an affibody-based affinity column, suitable for recovery of different variants of Z(HER2) affibody molecules, having a common binding surface for HER2. Eight such Z(HER2) affibody molecules, designed for future radioimaging investigations, having different C-terminal peptide extensions aimed for radioisotope ((??m)Tc)-chelation, were successfully produced and recovered in a single step to high purity using the anti-idiotypic affibody ligand for the affinity purification. These results clearly suggest a potential for the development of anti-idiotypic affibody-based resins for efficient recovery of related variants of a target protein that might have altered biochemical properties, thus avoiding the cumbersome design of specific recovery schemes for each variant of a target protein.     

Reconstructing past ecological networks: the reconfiguration of seed-dispersal interactions after megafaunal extinction

The late Quaternary megafaunal extinction impacted ecological communities worldwide, and affected key ecological processes such as seed dispersal. The traits of several species of large-seeded plants are thought to have evolved in response to interactions with extinct megafauna, but how these extinctions affected the organization of interactions in seed-dispersal systems is poorly understood. Here, we combined ecological and paleontological data and network analyses to investigate how the structure of a species-rich seed-dispersal network could have changed from the Pleistocene to the present and examine the possible consequences of such changes. Our results indicate that the seed-dispersal network was organized into modules across the different time periods but has been reconfigured in different ways over time. The episode of megafaunal extinction and the arrival of humans changed how seed dispersers were distributed among network modules. However, the recent introduction of livestock into the seed-dispersal system partially restored the original network organization by strengthening the modular configuration. Moreover, after megafaunal extinctions, introduced species and some smaller native mammals became key components for the structure of the seed-dispersal network. We hypothesize that such changes in network structure affected both animal and plant assemblages, potentially contributing to the shaping of modern ecological communities. The ongoing extinction of key large vertebrates will lead to a variety of context-dependent rearranged ecological networks, most certainly affecting ecological and evolutionary processes.     

Effect of tortuous extracellular pathways on resistance measurements

There are many instances in which we are limited to measuring macroscopic quantities such as a bulk flow or an average field. In biology, wer are frequently interested in using such macroscopic measurements, for example, the total current from a tissue, to determine the microscopic properties of the cells or tubules of the tissue. The microstructure of the tissue will generally increase the resistance to flow over what would be measured in an unstructured medium. This paper derives a fairly general expression for the relationship between effective resistance to macroscopic flow and the specific resistance of the medium conducting the microscopic flow. This expression, called a tortuosity factor, is defined entirely in terms of measurable morphometric and geometric parameters of the tissue.