Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes
Abstract The difficulties in purification of VLP-based recombinant hepatitis B surface antigen (rHBsAg) are mainly emerged from inefficient semi-purification step plus proteins physicochemical properties and these issues make the downstream processing (DSP) very lengthy and expensive. In this study,...
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| Format: | Article |
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Nature Portfolio
2023-07-01
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| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-023-37614-y |
| _version_ | 1797784621224558592 |
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| author | Maryam Moazami Goodarzi Reza Jalalirad Delaram Doroud Hamidreza Hozouri Mohammadreza Aghasadeghi Mahdi Paryan |
| author_facet | Maryam Moazami Goodarzi Reza Jalalirad Delaram Doroud Hamidreza Hozouri Mohammadreza Aghasadeghi Mahdi Paryan |
| author_sort | Maryam Moazami Goodarzi |
| collection | DOAJ |
| description | Abstract The difficulties in purification of VLP-based recombinant hepatitis B surface antigen (rHBsAg) are mainly emerged from inefficient semi-purification step plus proteins physicochemical properties and these issues make the downstream processing (DSP) very lengthy and expensive. In this study, optimization of rHBsAg (recombinantly-expressed in Pichia pastoris) DSP was performed using selection of buffering conditions in the semi-purification step. In the semi-purification optimization step, up to 73% of the protein impurities were eliminated and the utmost increase in rHBsAg purity (ca. 3.6-fold) was achieved using 20 mM sodium acetate, pH 4.5. By using rHBsAg binding and nonbinding situations obtained from the response surface plot in design of experiments (DOE), additional bind-elute and flow-through purification mode experiments were conducted and rHBsAg with high purity (near 100%) and recovery (> 83%) was achieved. Following assessment of critical quality attributes (i.e., purity, particle size distribution, host cell DNA, host cell protein, secondary structures, specific activity and relative potency), it was indicated that the characteristics of rHBsAg purified by the new DSP were similar or superior to the ones obtained from conventional DSP. The purification performance of the resin was constantly retained (97–100%) and no significant resin damage took place after 10 adsorption–elution–cleaning cycles. The new DSP developed for production of rHBsAg in this study can substitute the conventional one with granting satisfactory target protein quality, long-lasting resin efficacy, shorter and less expensive process. This process may be also employable for purification of both non-VLP- and VLP- based target proteins expressed in the yeast. |
| first_indexed | 2024-03-13T00:43:32Z |
| format | Article |
| id | doaj.art-5aefb3f6b7784a1480d4beb7e6262178 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-03-13T00:43:32Z |
| publishDate | 2023-07-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-5aefb3f6b7784a1480d4beb7e62621782023-07-09T11:12:06ZengNature PortfolioScientific Reports2045-23222023-07-0113112110.1038/s41598-023-37614-yDetermining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modesMaryam Moazami Goodarzi0Reza Jalalirad1Delaram Doroud2Hamidreza Hozouri3Mohammadreza Aghasadeghi4Mahdi Paryan5Department of Research and Development, Production and Research Complex, Pasteur Institute of IranDepartment of Research and Development, Production and Research Complex, Pasteur Institute of IranDepartment of Research and Development, Production and Research Complex, Pasteur Institute of IranDepartment of Quality Management, Production and Research Complex, Pasteur Institute of IranDepartment of Hepatitis and AIDS, Pasteur Institute of IranDepartment of Research and Development, Production and Research Complex, Pasteur Institute of IranAbstract The difficulties in purification of VLP-based recombinant hepatitis B surface antigen (rHBsAg) are mainly emerged from inefficient semi-purification step plus proteins physicochemical properties and these issues make the downstream processing (DSP) very lengthy and expensive. In this study, optimization of rHBsAg (recombinantly-expressed in Pichia pastoris) DSP was performed using selection of buffering conditions in the semi-purification step. In the semi-purification optimization step, up to 73% of the protein impurities were eliminated and the utmost increase in rHBsAg purity (ca. 3.6-fold) was achieved using 20 mM sodium acetate, pH 4.5. By using rHBsAg binding and nonbinding situations obtained from the response surface plot in design of experiments (DOE), additional bind-elute and flow-through purification mode experiments were conducted and rHBsAg with high purity (near 100%) and recovery (> 83%) was achieved. Following assessment of critical quality attributes (i.e., purity, particle size distribution, host cell DNA, host cell protein, secondary structures, specific activity and relative potency), it was indicated that the characteristics of rHBsAg purified by the new DSP were similar or superior to the ones obtained from conventional DSP. The purification performance of the resin was constantly retained (97–100%) and no significant resin damage took place after 10 adsorption–elution–cleaning cycles. The new DSP developed for production of rHBsAg in this study can substitute the conventional one with granting satisfactory target protein quality, long-lasting resin efficacy, shorter and less expensive process. This process may be also employable for purification of both non-VLP- and VLP- based target proteins expressed in the yeast.https://doi.org/10.1038/s41598-023-37614-y |
| spellingShingle | Maryam Moazami Goodarzi Reza Jalalirad Delaram Doroud Hamidreza Hozouri Mohammadreza Aghasadeghi Mahdi Paryan Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes Scientific Reports |
| title | Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes |
| title_full | Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes |
| title_fullStr | Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes |
| title_full_unstemmed | Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes |
| title_short | Determining buffer conditions for downstream processing of VLP-based recombinant hepatitis B surface antigen using multimodal resins in bind-elute and flow-through purification modes |
| title_sort | determining buffer conditions for downstream processing of vlp based recombinant hepatitis b surface antigen using multimodal resins in bind elute and flow through purification modes |
| url | https://doi.org/10.1038/s41598-023-37614-y |
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