Thermostable Peroxidase−Polylysine Films for Biocatalysis at 90 °C

Abstract

ross-linked films of poly(l-lysine) (PLL) and enzymes covalently linked to surfaces provided remarkable thermostability, enabling biocatalysis at 90 °C. Soret spectra, circular dichroism, and voltammetry showed that PLL films containing peroxidases or myoglobin were stable for up to 9 h at 90 °C, while the same enzymes in solution denatured completely within 20 min. Biocatalytic reduction of t-BuOOH with enzyme-PLL films, using rotating disk voltammetry, provided Michaelis kcat/Km values. Results showed that horseradish peroxidase (HRP)-PLL is 3-fold more active than soybean peroxidase (SBP)-PLL at 25 °C, but SBP-PLL is slightly more active at 90 °C. SBP-PLL films had 8-fold larger kcat/Km values at 90 °C compared to 25 °C. Oxidation of o-methoxyphenol to 3,3‘-dimethoxy-4,4‘-biphenoquinone by peroxidase-PLL-coated silica colloids gave better yields at 90 °C than 25 °C, suggesting increasing catalytic efficiency and selectivity at the higher temperature. These biocolloids were reusable with little loss of activity at 90 °C.