A novel heat-resistant hydroxycinnamic acid decarboxylase from Wickerhamomyces anomalus.

Hydroxycinnamate decarboxylase (HCDC) can decarboxylate Hydroxycinnamic acid to produce vinylphenol. In this study, a new heat-resistant HCDC (WaHCDC) was obtained from Wickerhamomyces anomalus WA64 isolated from Daqu, the fermentation starter of Chinese Baijiu. Subsequently, WaHCDC was expressed in P. pastoris GS115. After purification with Ni2+-NTA resin, WaHCDC was obtained at 25 mg·L-1 with a molecular weight of 45 kDa. The enzyme activity of WaHCDC was 12.69 ± 0.07 U·mg-1. The relative activity of WaHCDC was still retained 57 % after incubating at 80 °C for 90 min. To enhance acid resistance, WaHCDC was immobilized on covalent organic frameworks (COFs) via in-situ encapsulation, achieving 80 % loading efficiency and a capacity of 1.5 mg·g-1. The immobilized activity was 14.35 ± 0.11 U·mg-1. The stability of the immobilized WaHCDC was improved at pH 3-6. Its conversion rate of ferulic acid (FA) to 4-vinylguaiacol was 13 % higher than that of the free WaHCDC after incubating at 40 °C for 120 min. The activity of immobilized WaHCDC was above 60 % after 7 cycles reusing. This study provides a new heat-resistant enzyme resource and efficient immobilization strategy for the green biosynthesis of vinylphenol.