Owing to peculiar properties of nanobody, including nanoscale size, robust structure, stable and soluble behaviors in aqueous solution, reversible refolding, high affinity and specificity for only one cognate target, superior cryptic cleft accessibility, and deep tissue penetration, as well as a sustainable source, it has been an ideal research tool for the development of sophisticated nanobiotechnologies. Currently, the nanobody has been evolved into versatile research and application tool kits for diverse biomedical and biotechnology applications. Various nanobody-derived formats, including the nanobody itself, the radionuclide or fluorescent-labeled nanobodies, nanobody homo- or heteromultimers, nanobody-coated nanoparticles, and nanobody-displayed bacteriophages, have been successfully demonstrated as powerful nanobiotechnological tool kits for basic biomedical research, targeting drug delivery and therapy, disease diagnosis, bioimaging, and agricultural and plant protection. These applications indicate a special advantage of these nanobody-derived technologies, already surpassing the “me-too” products of other equivalent binders, such as the full-length antibodies, single-chain variable fragments, antigen-binding fragments, targeting peptides, and DNA-based aptamers. In this review, we summarize the current state of the art in nanobody research, focusing on the nanobody structural features, nanobody production approach, nanobody-derived nanobiotechnology tool kits, and the potentially diverse applications in biomedicine and biotechnology. The future trends, challenges, and limitations of the nanobody-derived nanobiotechnology tool kits are also discussed.

Phytases are widely utilized in feed industry to increase the utilization of phosphorus, minerals, and amino acids for improvement of animal and human nutrition. At present, all known β-propeller phytases (BPP) have been generated from bacteria, particularly Bacillus. In this work we report for the first time a new fungal-derived BPP phytase. We identified a phytase highly differentially expressed only in the parasitic stage of a nematophagous fungus, Arhtrobotrys oliogospora, during the development of the 3D traps. We found that this phytase was homologous to the known bacterial BPP phytase, thus we referred the new phytase to Aophytase. The heterologous expression of codon-optimized Aophytase gene in Pichia pastoris was successfully investigated to yield recombinant Aophytase (r-Aophytase) with high specific enzyme activity of 74.71 U/mg, much higher than those of recombinant BPP phytases derived bacteria. The kinetic parameters of the r-Aophytase, the optimum pH and temperature, as well as the effects of surfactant, EDTA and different ions on its enzyme activity were further investigated. The potential utilization of r-Aophytase in feed processing was finally explored. We found that the optimal pH value was about 7.5, and the optimal temperature was 50 °C.; r-Aophytase significantly increased the release of inorganic phosphorus from soybean meal, and improved the release of soluble minerals from the durum wheat flour and finger millet flour. The findings indicate its potential utilization in the feed processing to ameliorate nutritional value of cereals and animal feed in the future.