title: Optimization of protein electroextraction from microalgae by a flow process.
abstract: Classical methods, used for large scale treatments such as mechanical or chemical extractions, affect the integrity of extracted cytosolic protein by releasing proteases contained in vacuoles. Our previous experiments on flow processes electroextraction on yeasts proved that pulsed electric field technology allows preserving the integrity of released cytosolic proteins, by not affecting vacuole membranes. Furthermore, large cell culture volumes are easily treated by the flow technology. Based on this previous knowledge, we developed a new protocol in order to electro-extract total cytoplasmic proteins from microalgae (Nannochloropsis salina, Chlorella vulgaris and Haematococcus pluvialis). Given that induction of electropermeabilization is under the control of target cell size, as the mean diameter for N. salina is only 2.5 μm, we used repetitive 2 ms long pulses of alternating polarities with stronger field strengths than previously described for yeasts. The electric treatment was followed by a 24h incubation period in a salty buffer. The amount of total protein release was observed by a classical Bradford assay. A more accurate evaluation of protein release was obtained by SDS-PAGE. Similar results were obtained with C. vulgaris and H. pluvialis under milder electrical conditions as expected from their larger size.
authors.affiliation: CNRS UMR5089-IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 Route de Narbonne, BP 64182, F-31077 Toulouse, France; Université de Toulouse, UPS, IPBS, F-31077 Toulouse, France.
authors.forename: Mathilde
authors.initials: M
authors.lastname: Coustets
authors.affiliation: CERPEM, Maison de la Technopole, 6 rue Léonard de Vinci, F-53000 Laval, France.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Joubert-Durigneux
authors.affiliation: Sea, Molecules, Health (EA2160), IUML - FR 3473 CNRS, Université du Maine, IUT de Laval, Rue des Drs Calmette et Guerin, F-53020 Laval Cedex 9, France.
authors.forename: Josiane
authors.initials: J
authors.lastname: Hérault
authors.affiliation: MicroMar, Mer, Molécules, Santé, IUML - FR 3473 CNRS, University of Le Mans, Avenue Olivier Messiaen, F-72085 Le Mans cedex 9, France.
authors.forename: Benoît
authors.initials: B
authors.lastname: Schoefs
authors.affiliation: Sea, Molecules, Health (EA2160), IUML - FR 3473 CNRS, Université du Maine, IUT de Laval, Rue des Drs Calmette et Guerin, F-53020 Laval Cedex 9, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Blanckaert
authors.affiliation: CERPEM, Maison de la Technopole, 6 rue Léonard de Vinci, F-53000 Laval, France.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Garnier
authors.affiliation: CNRS UMR5089-IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 Route de Narbonne, BP 64182, F-31077 Toulouse, France; Université de Toulouse, UPS, IPBS, F-31077 Toulouse, France. Electronic address: justin.teissie@ipbs.fr.
authors.forename: Justin
authors.initials: J
authors.lastname: Teissié
mesh_terms: D005591:Chemical Fractionation / Q000379:methods*; D048408:Chlorella vulgaris / Q000737:chemistry; D000460:Chlorophyta / Q000737:chemistry; D003600:Cytosol / Q000737:chemistry; D055664:Electrochemical Techniques / Q000379:methods; D004586:Electrophoresis; D018274:Electroporation / Q000379:methods; D007700:Kinetics; D058086:Microalgae / Q000737:chemistry*; D010865:Pilot Projects; D011506:Proteins / Q000302:isolation & purification*; D013696:Temperature
chemical_list: D011506:Proteins
keywords: Electroextraction; Flow process; Microalgae; Pulsed electric field