title: Combined hydrophobic-metal binding fusion tags for applications in aqueous two-phase partitioning.
abstract: In this work, we studied the influence of fusion affinity tags containing both hydrophobic and histidines residues on the partitioning of the green fluorescent protein, GFPuv, in aqueous two-phase system. The tags were fused to the N-terminal of GFPuv and tested by immobilized metal affinity partitioning, in a PEG/salt system. The presence of both types of residues in the tag increased the partitioning greatly. Particularly, four engineered tags (H6, FH6, WH6, and YH6) containing a hexa-histidine sequence as well as different hydrophobic residues, all increased partitioning more than twice, reaching K values around 20, as compared to another construct (His6-GFP) containing an isolated hexa-histidine sequence. YH6, also proved be beneficial for protein expression.
authors.affiliation: Department of Pure and Applied Biochemistry, Centre for Chemistry and Chemical Engineering, Lund University, P.O. Box 124, 221 00 Lund, Sweden. florent.bernaudat@tbiokem.lth.se
authors.forename: Florent
authors.initials: F
authors.lastname: Bernaudat
authors.forename: Leif
authors.initials: L
authors.lastname: Bülow
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods; D006639:Histidine / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D057927:Hydrophobic and Hydrophilic Interactions; D008670:Metals / Q000737:chemistry; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D015202:Protein Engineering / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
chemical_list: C471213:His-His-His-His-His-His; D008670:Metals; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D006639:Histidine