title: Insertional-fusion of basic fibroblast growth factor endowed ribonuclease 1 with enhanced cytotoxicity by steric blockade of inhibitor interaction.
abstract: Basic fibroblast growth factor (bFGF) was inserted in the middle of human ribonuclease 1 (RNase1) sequence at an RNase inhibitor (RI)-binding site (Gly89) by a new gene fusion technique, insertional-fusion. The resultant insertional-fusion protein (CL-RFN89) was active both as bFGF and as RNase. Furthermore, it acquired an additional ability of evading RI through steric blockade of RI-binding caused by fused bFGF domain. As a result, CL-RFN89 showed stronger growth inhibition on B16/BL6 melanoma cells than an RI-sensitive tandem fusion protein. Thus, the insertional-fusion technique increases accessible positions for gene fusion on RNase, resulting in construction of a potent cytotoxic RNase.
authors.affiliation: Department of Bioscience and Biotechnology, Faculty of Engineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan. tadahrk@biotech.okayama-u.ac.jp
authors.forename: Hiroko
authors.initials: H
authors.lastname: Tada
authors.forename: Masayuki
authors.initials: M
authors.lastname: Onizuka
authors.forename: Kazuko
authors.initials: K
authors.lastname: Muraki
authors.forename: Wataru
authors.initials: W
authors.lastname: Masuzawa
authors.forename: Junichiro
authors.initials: J
authors.lastname: Futami
authors.forename: Megumi
authors.initials: M
authors.lastname: Kosaka
authors.forename: Masaharu
authors.initials: M
authors.lastname: Seno
authors.forename: Hidenori
authors.initials: H
authors.lastname: Yamada
mesh_terms: D000818:Animals; D001483:Base Sequence; D002455:Cell Division; D045744:Cell Line, Tumor; D017931:DNA Primers; D004791:Enzyme Inhibitors / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D004791:Enzyme Inhibitors; D011993:Recombinant Fusion Proteins; D016222:Fibroblast Growth Factor 2; D012259:Ribonuclease, Pancreatic