title: Grafting synthetic transmembrane units to the engineered low-toxicity α-hemolysin to restore its hemolytic activity.
abstract: The chemical modification of proteins to provide desirable functions and/or structures broadens their possibilities for use in various applications. Usually, proteins can acquire new functions and characteristics, in addition to their original ones, via the introduction of synthetic functional moieties. Here, we adopted a more radical approach to protein modification, i.e., the replacement of a functional domain of proteins with alternative chemical compounds to build "cyborg proteins." As a proof of concept model, we chose staphylococcal α-hemolysin (Hla), which is a well-studied, pore-forming toxin. The hemolytic activity of Hla mutants was dramatically decreased by truncation of the stem domain, which forms a β-barrel pore in the membrane. However, the impaired hemolytic activity was significantly restored by attaching a pyrenyl-maleimide unit to the cysteine residue that was introduced in the remaining stem domain. In contrast, negatively charged fluorescein-maleimide completely abolished the remaining activity of the mutants.
authors.affiliation: Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, 2-1-1, Katahira, Aoba-ku, Sendai 980-8577, Japan. kinbara@tagen.tohoku.ac.jp.
authors.forename: Mihoko
authors.initials: M
authors.lastname: Ui
authors.forename: Kousuke
authors.initials: K
authors.lastname: Harima
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Takei
authors.forename: Kouhei
authors.initials: K
authors.lastname: Tsumoto
authors.forename: Kazuhito V
authors.initials: KV
authors.lastname: Tabata
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Noji
authors.forename: Sumire
authors.initials: S
authors.lastname: Endo
authors.forename: Kimio
authors.initials: K
authors.lastname: Akiyama
authors.forename: Takahiro
authors.initials: T
authors.lastname: Muraoka
authors.forename: Kazushi
authors.initials: K
authors.lastname: Kinbara
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics; D003545:Cysteine / Q000737:chemistry; D004912:Erythrocytes; D005452:Fluoresceins / Q000737:chemistry; D005786:Gene Expression Regulation; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000235:genetics; D006461:Hemolysis; D006863:Hydrogen-Ion Concentration; D008301:Maleimides / Q000737:chemistry; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D012756:Sheep; D013211:Staphylococcus aureus
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D005452:Fluoresceins; D006460:Hemolysin Proteins; D008301:Maleimides; C021091:staphylococcal alpha-toxin; C043592:maleimide; C037041:fluorescein 5-maleimide; D003545:Cysteine