A new insight in gellan microspheres application to capture a plasmid DNA vaccine from an Escherichia coli lysate

Gomes, Diana; Costa, Diana; Queiroz, João A.; Passarinha, Luís A.; Sousa, Ângela

doi:10.1016/j.seppur.2021.119013

Cited by 5 publications

(4 citation statements)

References 44 publications

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“…This phenomenon highlights the effect that inherent biomolecule properties can have on establishing biointeractions with GG microspheres. For instance, our research group has previously relied on similar GG microsphere batch methods as the primary purification technique for the purification of soluble proteins [35] and plasmid DNA [36], without the rise of any complexation issues.…”

Section: Discussionmentioning

confidence: 99%

“…Nickel-, magnesium-and copper-crosslinked GG microspheres had also been previously reported with a spherical shape. However, the aforementioned microspheres presented clear rugosity, cavities and irregularities in their surface [35,36]. By comparison, calcium-crosslinked GG microspheres are far smoother, with no apparent pores, cavities or cracks (Figure 1D).…”

Section: Characterization Of Gellan Gum Microspheresmentioning

confidence: 96%

“…When compared to nickel-crosslinked GG microspheres, the normalized ion percentage levels seem to differ nearly two-fold. Indeed, when comparing copper-, magnesium-, nickel-and calcium-crosslinked microspheres by crosslinker percentage, it seems that transition metals are incorporated in higher degrees than alkaline earth metals [35,36]. This stronger crosslinker concentration can induce the formation of a tighter mesh network, resulting in more compact microspheres, which can justify why nickel-crosslinked microspheres are smaller than calcium-crosslinked microspheres [45].…”

Section: Characterization Of Gellan Gum Microspheresmentioning

confidence: 99%

“…Indeed, GG-based materials have been shown to promote strong adsorption of small drug molecules [34]. Recently, our research group demonstrated that GG microspheres can efficiently capture soluble catechol-O-methyltransferase (COMT) [35] and plasmid DNA [36]. Current microsphere-based methods being developed for protein capture are mostly based on magnetic microspheres, which quickly enhance the complexity and cost of bioprocesses [37][38][39][40].…”

Section: Introductionmentioning

confidence: 99%

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Specific Six-Transmembrane Epithelial Antigen of the Prostate 1 Capture with Gellan Gum Microspheres: Design, Optimization and Integration

Batista-Silva

Gomes

Barroca-Ferreira

et al. 2023

IJMS

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This work demonstrates the potential of calcium- and nickel-crosslinked Gellan Gum (GG) microspheres to capture the Six-Transmembrane Epithelial Antigen of the Prostate 1 (STEAP1) directly from complex Komagataella pastoris mini-bioreactor lysates in a batch method. Calcium-crosslinked microspheres were applied in an ionic exchange strategy, by manipulation of pH and ionic strength, whereas nickel-crosslinked microspheres were applied in an affinity strategy, mirroring a standard immobilized metal affinity chromatography. Both formulations presented small diameters, with appreciable crosslinker content, but calcium-crosslinked microspheres were far smoother. The most promising results were obtained for the ionic strategy, wherein calcium-crosslinked GG microspheres were able to completely bind 0.1% (v/v) DM solubilized STEAP1 in lysate samples (~7 mg/mL). The target protein was eluted in a complexed state at pH 11 with 500 mM NaCl in 10 mM Tris buffer, in a single step with minimal losses. Coupling the batch clarified sample with a co-immunoprecipitation polishing step yields a sample of monomeric STEAP1 with a high degree of purity. For the first time, we demonstrate the potential of a gellan batch method to function as a clarification and primary capture method towards STEAP1, a membrane protein, simplifying and reducing the costs of standard purification workflows.

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Section: Discussionmentioning

confidence: 99%

Section: Characterization Of Gellan Gum Microspheresmentioning

confidence: 96%