Hyaluronidases are a group of glycosidases catalyzing the degradation of hyaluronic acid (HA). Because of the advantages of effectively hydrolyzing the HA-rich matrix and low immunogenicity, human hyaluronidase PH20 (hPH20) is widely used in the medical field. Here, we realized the active expression of recombinant hPH20 by Pichia pastoris under a methanol-induced promoter PAOX1. By optimizing the composition of the C-terminal domain and fusing protein tags, we constructed a fusion mutant AP2-△491C with the extracellular hyaluronidase activity of 258.1 U·L−1 in a 3-L bioreactor, the highest expression level of recombinant hPH20 produced by microbes. Furthermore, we found recombinant hPH20 hydrolyzed the β-1,4 glycosidic bonds sequentially from the reducing end of o-HAs, with HA6NA as the smallest substrate. The result will provide important theoretical guidance for the directed evolution of the enzyme to prepare multifunctional o-HAs with specific molecular weights.
CITATION STYLE
Pang, B., He, J., Zhang, W., Huang, H., Wang, Y., Wang, M., … Kang, Z. (2022). Active Expression of Human Hyaluronidase PH20 and Characterization of Its Hydrolysis Pattern. Frontiers in Bioengineering and Biotechnology, 10. https://doi.org/10.3389/fbioe.2022.885888
Mendeley helps you to discover research relevant for your work.