Purification of Recombinant Human FGF1 and Mutant

Wild-type human FGF1 and mutant FGF1 (FGF1-PIGN), shown in Fig. 1A, were purified as described previously [15 (link), 16 (link)]. Briefly, mutations of the human FGF1 gene were introduced using the QuikChange site-directed mutagenesis kit (Agilent Technologies, Santa Clara, CA). The wild-type FGF1 or FGF1-PIGN gene was transferred into the pDEST17 vector (Thermo Fisher Scientific, Waltham, MA), an N-terminal fusion vector that contains a sequence encoding a 6 × His tag, and the pDEST17 expression constructs were transformed into BL21(DE3)pLysS Escherichia coli cells. Protein expression induction was performed using the Overnight Express Autoinduction System 1 (Merck kGaA, Darmstadt, Germany), according to the manufacturer’s instructions. The cell pellets were lysed in BugBuster Master Mix (Merck kGaA) including ethylenediaminetetraacetic acid (EDTA)-free protease inhibitor cocktail (cOmplete ULTRA) (Roche Diagnostics, Mannheim, Germany), and soluble extracts purified using a Ni Sepharose High Performance column (GE Healthcare, Waukesha, WI).