Development of a biosensing platform based on a laccase-hydrophobin chimera

Appl Microbiol Biotechnol. 2019 Apr;103(7):3061-3071. doi: 10.1007/s00253-019-09678-2. Epub 2019 Feb 19.


A simple and stable immobilization of a laccase from Pleurotus ostreatus was obtained through genetic fusion with a self-assembling and adhesive class I hydrophobin. The chimera protein was expressed in Pichia pastoris and secreted into the culture medium. The crude culture supernatant was directly used for coatings of polystyrene multi-well plates without additional treatments, a procedure that resulted in a less time-consuming and chemicals reduction. Furthermore, the gene fusion yielded a positive effect with respect to the wild-type recombinant enzyme in terms of both immobilization and stability. The multi-well plate with the immobilized chimera was used to develop an optical biosensor to monitor two phenolic compounds: L-DOPA ((S)-2-amino-3-(3,4-dihydroxyphenyl) propanoic acid) and caffeic acid (3-(3,4-dihydroxyphenyl)-2-propenoic acid); the estimation of which is a matter of interest in the pharmaceutics and food field. The method was based on the use of the analytes as competing inhibitors of the laccase-mediated ABTS oxidation. The main advantages of the developed biosensor are the ease of preparation, the use of small sample volumes, and the simultaneous analysis of multiple samples on a single platform.

Keywords: Caffeic acid; Chimera proteins; Immobilization; L-DOPA; Self-assembly.

MeSH terms

  • Biosensing Techniques*
  • Caffeic Acids / metabolism
  • Cloning, Molecular
  • Culture Media / chemistry
  • Enzymes, Immobilized / biosynthesis
  • Fungal Proteins / biosynthesis*
  • Fungal Proteins / genetics
  • Hydrogen-Ion Concentration
  • Laccase / biosynthesis*
  • Laccase / genetics
  • Levodopa / metabolism
  • Oxidation-Reduction
  • Pichia / genetics
  • Pleurotus / enzymology*
  • Polystyrenes
  • Recombinant Fusion Proteins / biosynthesis


  • Caffeic Acids
  • Culture Media
  • Enzymes, Immobilized
  • Fungal Proteins
  • Polystyrenes
  • Recombinant Fusion Proteins
  • Levodopa
  • Laccase
  • caffeic acid