title: Lactococcus lactis is an Efficient Expression System for Mammalian Membrane Proteins Involved in Liver Detoxification, CYP3A4, and MGST1.
abstract: Despite the great importance of human membrane proteins involved in detoxification mechanisms, their wide use for biochemical approaches is still hampered by several technical difficulties considering eukaryotic protein expression in order to obtain the large amounts of protein required for functional and/or structural studies. Lactococcus lactis has emerged recently as an alternative heterologous expression system to Escherichia coli for proteins that are difficult to express. The aim of this work was to check its ability to express mammalian membrane proteins involved in liver detoxification, i.e., CYP3A4 and two isoforms of MGST1 (rat and human). Genes were cloned using two different strategies, i.e., classical or Gateway-compatible cloning, and we checked the possible influence of two affinity tags (6×-His-tag and Strep-tag II). Interestingly, all proteins could be successfully expressed in L. lactis at higher yields than those previously obtained for these proteins with classical expression systems (E. coli, Saccharomyces cerevisiae) or those of other eukaryotic membrane proteins expressed in L. lactis. In addition, rMGST1 was fairly active after expression in L. lactis. This study highlights L. lactis as an attractive system for efficient expression of mammalian detoxification membrane proteins at levels compatible with further functional and structural studies.
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Sana
authors.initials: S
authors.lastname: Bakari
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Lembrouk
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Laura
authors.initials: L
authors.lastname: Sourd
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Fares
authors.initials: F
authors.lastname: Ousalem
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: François
authors.initials: F
authors.lastname: André
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Orlowski
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France.
authors.forename: Marcel
authors.initials: M
authors.lastname: Delaforge
authors.affiliation: Institute of Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ. Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette Cedex, France. annie.frelet-barrand@femto-st.fr.|Institute FEMTO-ST, UMR6174 CNRS-Université de Franche-Comté, 25044, Besançon Cedex, France. annie.frelet-barrand@femto-st.fr.
authors.forename: Annie
authors.initials: A
authors.lastname: Frelet-Barrand
mesh_terms: D000818:Animals; D003001:Cloning, Molecular / Q000379:methods*; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D013294:Lactococcus lactis / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism
chemical_list: D008565:Membrane Proteins; D011994:Recombinant Proteins; D051544:Cytochrome P-450 CYP3A; C122107:microsomal glutathione S-transferase-I; D005982:Glutathione Transferase
keywords: Cytochrome P450; Gateway; Lactococcus lactis; Membrane proteins; Microsomal glutathione S-transferase