Applications of gellan natural polymer microspheres in recombinant catechol-O-methyltransferase direct capture from a Komagataella pastoris lysate

Abstract

The present work shows the application of nickel- and magnesium-crosslinked gellan microspheres in ionic and affinity capture strategies to directly extract hSCOMT from the complex Komagataella pastoris lysate through a simple batch method. Both formulations present similar morphology, but nickel-crosslinked microspheres present higher crosslinker content and smaller diameters. Four different capture strategies were established, by manipulating the ionic strength, pH, temperature and competing agents' presence. The most promising results for hSCOMT capture and clarification were obtained employing an ionic strategy with nickel-crosslinked microspheres and an affinity strategy with magnesium-crosslinked microspheres at 4 °C. The bioactivity results (200%) and purification degree (70%) of hSCOMT captured by the ionic strategy were more satisfactory probably due to the soft ionic conditions used (100 mM NaCl). For the first time, the gellan polysaccharide versatility was demonstrated in the microsphere application for the direct capture of hSCOMT from a complex lysate, simplifying isolation biotechnological procedures.

Keywords: Affinity interactions; Batch method; Capture strategies; Gellan microspheres; Human soluble catechol-O-methyltransferase; Ionic interactions.