title: Hydrophobin signal sequence mediates efficient secretion of recombinant proteins in Pichia pastoris.
abstract: Pichia pastoris is an important eukaryotic organism for the expression, processing, and secretion of recombinant proteins. Here, the secretion of enhanced green fluorescent protein (EGFP) in P. pastoris by using three novel secretion signals originating from the HFBI and HFBII class 2 hydrophobins of Trichoderma reesei was investigated. EGFP was fused to the carboxyl terminus of hydrophobin secretion signals and expressed under the control of the constitutive GAP promoter. In every case, recombinant EGFP entered the secretory pathway of P. pastoris. SDS-polyacrylamide gel electrophoresis, Western blot analysis of the cells' supernatant, and fluorescence measurements on single-cell level via flow cytometry confirmed the efficient secretion of EGFP mediated by the novel secretion sequences. In conclusion, the data clearly show that the secretion sequences derived from HFBI and HFBII of T. reesei have the potential to achieve an efficient secretion of heterologous proteins in P. pastoris. Due to the small size of the hydrophobin-derived secretion signals, their coding sequence can be easily introduced to the gene of interest by PCR.
authors.affiliation: Institute of Genetics, Dresden University of Technology, Zellescher Weg 20b, Dresden, Germany. Kirsten.Kottmeier@tu-dresden.de
authors.forename: Kirsten
authors.initials: K
authors.lastname: Kottmeier
authors.forename: Kai
authors.initials: K
authors.lastname: Ostermann
authors.forename: Thomas
authors.initials: T
authors.lastname: Bley
authors.forename: Gerhard
authors.initials: G
authors.lastname: Rödel
mesh_terms: D005110:Extracellular Space / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics*; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals; D021381:Protein Transport; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014242:Trichoderma / Q000235:genetics*
chemical_list: D005656:Fungal Proteins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins