title: Structure-function relationships in hydrophobins: probing the role of charged side chains.
abstract: Hydrophobins are small fungal proteins that are amphiphilic and have a strong tendency to assemble at interfaces. By taking advantage of this property, hydrophobins have been used for a number of applications: as affinity tags in protein purification, for protein immobilization, such as in foam stabilizers, and as dispersion agents for insoluble drug molecules. Here, we used site-directed mutagenesis to gain an understanding of the molecular basis of their properties. We especially focused on the role of charged amino acids in the structure of hydrophobins. For this purpose, fusion proteins consisting of Trichoderma reesei hydrophobin I (HFBI) and the green fluorescent protein (GFP) that contained various combinations of substitutions of charged amino acids (D30, K32, D40, D43, R45, K50) in the HFBI structure were produced. The effects of the introduced mutations on binding, oligomerization, and partitioning were characterized in an aqueous two-phase system. It was found that some substitutions caused better surface binding and reduced oligomerization, while some showed the opposite effects. However, all mutations decreased partitioning in surfactant systems, indicating that the different functions are not directly correlated and that partitioning is dependent on finely tuned properties of hydrophobins. This work shows that not all functions in self-assembly are connected in a predictable way and that a simple surfactant model for hydrophobin function is insufficient.
authors.affiliation: VTT Technical Research Centre of Finland, Espoo, Finland.
authors.forename: Michael
authors.initials: M
authors.lastname: Lienemann
authors.forename: Julie-Anne
authors.initials: JA
authors.lastname: Gandier
authors.forename: Jussi J
authors.initials: JJ
authors.lastname: Joensuu
authors.forename: Atsushi
authors.initials: A
authors.lastname: Iwanaga
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Takatsuji
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Haruyama
authors.forename: Emma
authors.initials: E
authors.lastname: Master
authors.forename: Maija
authors.initials: M
authors.lastname: Tenkanen
authors.forename: Markus B
authors.initials: MB
authors.lastname: Linder
mesh_terms: D019943:Amino Acid Substitution; D019677:Artificial Gene Fusion; D004252:DNA Mutational Analysis; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014242:Trichoderma / Q000235:genetics* / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins