title: In Vitro Comparison of Adipokine Export Signals.
abstract: Mammalian cells are widely used for recombinant protein production in research and biotechnology. Utilization of export signals significantly facilitates production and purification processes. 35 years after the discovery of the mammalian export machinery, there still are obscurities regarding the efficiency of the export signals. The aim of this study was the comparative evaluation of the efficiency of selected export signals using adipocytes as a cell model. Adipocytes have a large capacity for protein secretion including several enzymes, adipokines, and other signaling molecules, providing a valid system for a quantitative evaluation. Constructs that expressed N-terminal fusion export signals were generated to express Enhanced Green Fluorescence Protein (EGFP) as a reporter for quantitative and qualitative evaluation. Furthermore, fluorescent microscopy was used to trace the intracellular traffic of the reporter. The export efficiency of six selected proteins secreted from adipocytes was evaluated. Quantitative comparison of intracellular and exported fractions of the recombinant constructs demonstrated a similar efficiency among the studied sequences with minor variations. The export signal of Retinol Binding Protein (RBP4) exhibited the highest efficiency. This study presents the first quantitative data showing variations among export signals, in adipocytes which will help optimization of recombinant protein distribution.
authors.forename: Parisa
authors.initials: P
authors.lastname: Sharafi
authors.affiliation: Department of Medical Biology, Faculty of Medicine, Hacettepe University, Ankara, Turkey. kocaefe@hacettepe.edu.tr.
authors.forename: Y Çetin
authors.initials: YÇ
authors.lastname: Kocaefe
mesh_terms: D054392:Adipokines / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008297:Male; D021382:Protein Sorting Signals / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology*; D051381:Rats; D054839:Retinol-Binding Proteins, Plasma / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D054392:Adipokines; D021382:Protein Sorting Signals; C532035:Rbp4 protein, rat; D054839:Retinol-Binding Proteins, Plasma; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins