Urate oxidase purification by salting-in crystallization: towards an alternative to chromatography
| Autor: | Natalie Ferté, Mohamed El Hajji, Bertrand Castro, François Ragot, Marion Giffard, Françoise Bonneté |
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| Přispěvatelé: | Centre Interdisciplinaire de Nanoscience de Marseille (CINaM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Sanofi-Aventis R&D, SANOFI Recherche, Institut des Biomolécules Max Mousseron [Pôle Chimie Balard] (IBMM), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université de Montpellier (UM)-Ecole Nationale Supérieure de Chimie de Montpellier (ENSCM) |
| Jazyk: | angličtina |
| Rok vydání: | 2011 |
| Předmět: |
Macromolecular Assemblies
Urate Oxidase Polymers Cancer Treatment [CHIM.THER]Chemical Sciences/Medicinal Chemistry 02 engineering and technology Biochemistry Physical Chemistry law.invention chemistry.chemical_compound Engineering law Basic Cancer Research Biological Systems Engineering Rasburicase Materials Design Crystallization Solubility ComputingMilieux_MISCELLANEOUS chemistry.chemical_classification Chromatography 0303 health sciences Multidisciplinary Enzyme Classes Applied Chemistry Urate oxidase [SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences 021001 nanoscience & nanotechnology Enzymes Chemistry Physicochemical Properties Oncology Nephrology Chromatography Gel Salting out Medicine 0210 nano-technology Physical Organic Chemistry Research Article medicine.drug Science Materials Science Biophysics Salt (chemistry) Bioengineering Crystals 03 medical and health sciences [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication Chromatographic techniques medicine [CHIM]Chemical Sciences Crystallization techniques Biology 030304 developmental biology Chemotherapy and Drug Treatment Chemical Properties chemistry Fermentation Uric acid Salts Isoelectric Focusing Dialysis |
| Zdroj: | PLoS ONE, Vol 6, Iss 5, p e19013 (2011) PLoS ONE PLoS ONE, Public Library of Science, 2011, 6 (5), pp.e19013. ⟨10.1371/journal.pone.0019013⟩ PLoS ONE, Public Library of Science, 2011, ⟨10.1371/journal.pone.0019013⟩ PLoS ONE, 2011, 6 (5), pp.e19013. ⟨10.1371/journal.pone.0019013⟩ |
| ISSN: | 1932-6203 |
| DOI: | 10.1371/journal.pone.0019013⟩ |
| Popis: | BackgroundRasburicase (Fasturtec® or Elitek®, Sanofi-Aventis), the recombinant form of urate oxidase from Aspergillus flavus, is a therapeutic enzyme used to prevent or decrease the high levels of uric acid in blood that can occur as a result of chemotherapy. It is produced by Sanofi-Aventis and currently purified via several standard steps of chromatography. This work explores the feasibility of replacing one or more chromatography steps in the downstream process by a crystallization step. It compares the efficacy of two crystallization techniques that have proven successful on pure urate oxidase, testing them on impure urate oxidase solutions.Methodology/principal findingsHere we investigate the possibility of purifying urate oxidase directly by crystallization from the fermentation broth. Based on attractive interaction potentials which are known to drive urate oxidase crystallization, two crystallization routes are compared: a) by increased polymer concentration, which induces a depletion attraction and b) by decreased salt concentration, which induces attractive interactions via a salting-in effect. We observe that adding polymer, a very efficient way to crystallize pure urate oxidase through the depletion effect, is not an efficient way to grow crystals from impure solution. On the other hand, we show that dialysis, which decreases salt concentration through its strong salting-in effect, makes purification of urate oxidase from the fermentation broth possible.ConclusionsThe aim of this study is to compare purification efficacy of two crystallization methods. Our findings show that crystallization of urate oxidase from the fermentation broth provides purity comparable to what can be achieved with one chromatography step. This suggests that, in the case of urate oxidase, crystallization could be implemented not only for polishing or concentration during the last steps of purification, but also as an initial capture step, with minimal changes to the current process. |
| Databáze: | OpenAIRE |
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