The　fusion　of　cell　permeable　peptide　TAT　and　bifunctional　antioxidant　enzymes,　GST　(Glutathione　sulfur　transferase)-TAT-SOD1　(Cu,　Zn　superoxide　dismutase),　is　an　intracellular　superoxide　scavenger.　Compared　with　SOD1-TAT,　GST-TAT-SOD1　has　better　protective　effect　on　oxidative　damage　but　less　transduction　efficiency.　A　novel　cell　permeable　bifunctional　antioxidant　enzymes　with　the　fusion　of　GST,　SOD1　and　polyarginine　R9　was　constructed　for　higher　transduction　efficiency.　The　full　nucleotide　sequence　of　SOD1-R9　was　synthesized　and　inserted　into　the　prokaryotic　expression　vector　pGEX-4T-1　with　the　GST　tag.　After　the　successful　construction　of　the　prokaryotic　expression　vectors　of　GST-SOD1-R9,　the　recombinant　vector　was　then　transformed　into　Escherichia　coli　BL21　(DE3)　and　the　GST-SOD1-R9　fusion　protein　was　produced　with　the　induction　of　IPTG.　The　soluble　expression　of　GST-SOD1-R9　fusion　protein　was　combining　with　the　induction　temperature　and　time.　The　best　soluble　expression　was　obtained　with　the　induction　temperature　of　25　℃　and　the　induction　time　of　11　h.　The　fusion　protein　was　purified　through　the　combination　of　80%　ammonium　sulfate　precipitation　and　affinity　chromatography　using　glutathione　agarose,　and　verified　by　SDS-PAGE　and　special　enzymatic　activity.　The　thermal　and　pH　stability　of　GST-SOD1-R9　fusion　protein　were　analyzed　and　the　SOD　and　GST　activity　of　fusion　protein　were　proved　to　be　well　maintained　under　physiological　conditions.　Finally,　the　transduction　efficiency　of　GST-SOD1-R9　fusion　protein　was　proved　to　be　better　than　GST-TAT-SOD1　fusion　protein　(P＜0.05).　These　works　establish　a　foundation　for　further　study　of　the　protective　effect　of　GST-SOD1-R9　fusion　protein　against　oxidative　damage.　Keywords　©　2017,　Science　Press.　All　right　reserved.