Nitrilotriacetic Acid Functionalized Microgels for Efficient Immobilization of Hyaluronan Synthase

Macromolecular Bioscience, vol. 24, pp. e2400075

Abstract

Abstract Enzymes play a vital role in synthesizing complex biological molecules like hyaluronic acid (HA). Immobilizing enzymes on support materials is essential for their efficient use and reuse in multiple cycles. Microgels, composed of cross–linked, highly swollen polymer networks, are ideal for enzyme uptake owing to their high porosity. This study demonstrates the immobilization of His6‐tagged hyaluronan synthase from Pasteurella multocida (PmHAS) onto nitrilotriacetic acid functionalized microgels using different bivalent ions (Ni2+, Co2+, Mn2+, Mg2+, and Fe2+) via metal affinity binding. The results indicate that using Ni2+ yields the microgels with the highest enzyme uptake and HA formation. The immobilized PmHAS enables repetitive enzymatic production, producing high molecular weight HAs with decreasing dispersities in each step. Furthermore, the highest reported yield of HA with high molecular weight for immobilized PmHAS is achieved. This system establishes a foundation for continuous HA formation, with future works potentially enhancing PmHAS stability through protein engineering.

Authors 4

1. Isabel Katja Sommerfeld Aachen Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry

   DWI – Leibniz Institute for Interactive Materials · RWTH Aachen University

   Affiliation as printed

   DWI − Leibniz‐Institute for Interactive Materials e.V. Forckenbeckstraße 50 52074 Aachen Germany

   Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry RWTH Aachen University Worringerweg 2 52074 Aachen Germany

   Functional and Interactive Polymers, Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Worringerweg 2, 52074 Aachen, Germany

2. Esther Maria Dälken Aachen Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry

   DWI – Leibniz Institute for Interactive Materials · RWTH Aachen University

   Affiliation as printed

   DWI − Leibniz‐Institute for Interactive Materials e.V. Forckenbeckstraße 50 52074 Aachen Germany

   Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry RWTH Aachen University Worringerweg 2 52074 Aachen Germany

   Functional and Interactive Polymers, Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Worringerweg 2, 52074 Aachen, Germany

3. Lothar Elling corresponding Aachen

   RWTH Aachen University

   Affiliation as printed

   Laboratory for Biomaterials Institute of Biotechnology and Helmholtz‐Institute for Biomedical Engineering RWTH Aachen University Pauwelsstraße 20 52074 Aachen Germany

4. Andrij Pich corresponding Aachen Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry

   Maastricht University · DWI – Leibniz Institute for Interactive Materials · RWTH Aachen University

   Affiliation as printed

   Aachen Maastricht Institute for Biobased Materials (AMIBM) Maastricht University Brightlands Chemelot Campus Urmonderbaan 22 RD Geleen 6167 The Netherlands

   DWI − Leibniz‐Institute for Interactive Materials e.V. Forckenbeckstraße 50 52074 Aachen Germany

   Functional and Interactive Polymers Institute of Technical and Macromolecular Chemistry RWTH Aachen University Worringerweg 2 52074 Aachen Germany

   Aachen Maastricht Institute for Biobased Materials (AMIBM), Maastricht University, Brightlands Chemelot Campus, Urmonderbaan 22, RD Geleen, 6167 The Netherlands

   Functional and Interactive Polymers, Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Worringerweg 2, 52074 Aachen, Germany