title: Robust expression of a bioactive mammalian protein in Chlamydomonas chloroplast.
abstract: We have engineered the chloroplast of eukaryotic algae to produce a number of recombinant proteins, including human monoclonal antibodies, but, to date, have achieved expression to only 0.5% of total protein. Here, we show that, by engineering the mammalian coding region of bovine mammary-associated serum amyloid (M-SAA) as a direct replacement for the chloroplast psbA coding region, we can achieve expression of recombinant protein above 5% of total protein. Chloroplast-expressed M-SAA accumulates predominantly as a soluble protein, contains the correct amino terminal sequence and has little or no post-translational modification. M-SAA is found in mammalian colostrum and stimulates the production of mucin in the gut, acting in the prophylaxis of bacterial and viral infections. Chloroplast-expressed and purified M-SAA is able to stimulate mucin production in human gut epithelial cell lines. As Chlamydomonas reinhardtii is an edible alga, production of therapeutic proteins in this organism offers the potential for oral delivery of gut-active proteins, such as M-SAA.
authors.affiliation: The Department of Cell Biology and The Skaggs Institute for Chemical Biology, La Jolla, CA 92037, USA.
authors.forename: Andrea L
authors.initials: AL
authors.lastname: Manuell
authors.forename: Maria Verónica
authors.initials: MV
authors.lastname: Beligni
authors.forename: John H
authors.initials: JH
authors.lastname: Elder
authors.forename: David T
authors.initials: DT
authors.lastname: Siefker
authors.forename: Miller
authors.initials: M
authors.lastname: Tran
authors.forename: Annika
authors.initials: A
authors.lastname: Weber
authors.forename: Thomas L
authors.initials: TL
authors.lastname: McDonald
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Mayfield
mesh_terms: D020418:Algal Proteins / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D016825:Chlamydomonas reinhardtii / Q000235:genetics* / Q000378:metabolism; D002736:Chloroplasts / Q000235:genetics* / Q000378:metabolism; D018503:Genome, Protozoan; D030781:Organisms, Genetically Modified / Q000235:genetics / Q000378:metabolism; D010788:Photosynthesis; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D000685:Serum Amyloid A Protein / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D020418:Algal Proteins; D011994:Recombinant Proteins; D000685:Serum Amyloid A Protein