title: Purification and immunodetection of the complete recombinant HER-2[neu] receptor produced in yeast.
abstract: For the first time, the full length recombinant HER-2[neu] receptor has been produced in a yeast (Arxula adeninivorans). It is one of the most studied membrane receptors in oncology and is involved in aggressive tumor formation. A yeast integration rDNA cassette containing the human gene coding for the HER-2[neu] protein was constructed and a screening procedure was performed to select the most productive transformant. Different detergents were tested for efficient solubilization of the membrane bound protein, with CHAPS giving the best results. To increase the yield of the recombinant protein from HER-2[neu] producing A. adeninivorans, optimal culture parameters were established for cultivation in bioreactor. The recombinant protein was subsequently assayed using ELISA and SPR immunoassays systems with antibodies raised against two different epitopes of the human receptor. In both cases, elution fractions containing the recombinant HER-2[neu] receptor successfully reacted with the immunoassays with limits of quantification below 100ngml(-1). These results demonstrate that the full length recombinant HER-2[neu] reported here has the potential to be a new standard for the detection of HER-2 type cancer.
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Chamas
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Giersberg
authors.affiliation: Universitätsklinikum "Carl Gustav Carus" Dresden, Institut für Pathologie, Fetscherstr. 74, 01307 Dresden, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Friedrich
authors.affiliation: Fraunhofer Institut for Material and Beam Technology (IWS), Winterbergstr. 28, D-01277 Dresden, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Sonntag
authors.affiliation: Universitätsklinikum "Carl Gustav Carus" Dresden, Institut für Pathologie, Fetscherstr. 74, 01307 Dresden, Germany.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Kunze
authors.affiliation: quo data GmbH, Kaitzer Str. 135, D-01187 Dresden, Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Uhlig
authors.affiliation: new diagnostics GmbH, Moosstr. 92c, D-85356 Freising, Germany.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Simon
authors.affiliation: School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand.
authors.forename: Keith
authors.initials: K
authors.lastname: Baronian
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany. Electronic address: kunzeg@ipk-gatersleben.de.
authors.forename: Gotthard
authors.initials: G
authors.lastname: Kunze
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004718:Saccharomycetales / Q000235:genetics*; D020349:Surface Plasmon Resonance
chemical_list: D018719:Erb-b2 Receptor Tyrosine Kinases; D011994:Recombinant Proteins; C508053:ERBB2 protein, human
keywords: A. adeninivorans; Breast cancer; HER-2[neu]; Immunoassay; Protein solubilization; Surface Plasmon Resonance