Immobilization of a Novel ESTBAS Esterase from Bacillus altitudinis onto an Epoxy Resin: Characterization and Regioselective Synthesis of Chloramphenicol Palmitate

Wei Wei,Dongzhi Wei,Fengying Dong,Xiaohui Yang,Lin Lin,Dannong He,Xudong Tang

doi:10.3390/catal9070620

Abstract

Novel gene estBAS from Bacillus altitudinis, encoding a 216-amino acid esterase (EstBAS) with a signal peptide (SP), was expressed in Escherichia coli. EstBASΔSP showed the highest activity toward p-nitrophenyl hexanoate at 50 °C and pH 8.0 and had a half-life (T1/2) of 6 h at 50 °C. EstBASΔSP was immobilized onto a novel epoxy resin (Lx-105s) with a high loading of 96 mg/g. Fourier transform infrared (FTIR) spectroscopy showed that EstBASΔSP was successfully immobilized onto Lx-105s. In addition, immobilization improved its enzymatic performance by widening the tolerable ranges of pH and temperature. The optimum temperature of immobilized EstBASΔSP (Lx-EstBASΔSP) was higher, 60 °C, and overall thermostability improved. T1/2 of Lx-EstBASΔSP and free EstBASΔSP at 60 °C was 105 and 28 min, respectively. Lx-EstBASΔSP was used as a biocatalyst to synthesize chloramphenicol palmitate by regioselective modification at the primary hydroxyl group. Conversion efficiency reached 94.7% at 0.15 M substrate concentration after 24 h. Lx-EstBASΔSP was stable and could be reused for seven cycles, after which it retained over 80% of the original activity.

Highlights

Chloramphenicol was first isolated and purified from Streptomyces venezuelae by Burkholder in 1947 [1]
EstBAS and estBASΔSP from B. altitudinis were amplified with primer pairs F1/R1 and F2/R1, estBAS and fromframe
The open reading frame of est was found to consist of bp, encoding a protein of 216 amino acid residues with a molecular mass of 22.96 kDa and a theoretical pI of 9.85, as calculated amino acid residues with a molecular mass of kDa and a theoretical pI

Summary

Introduction

Chloramphenicol was first isolated and purified from Streptomyces venezuelae by Burkholder in 1947 [1]. Chloramphenicol was synthesized via chemical methods and produced on an industrial scale, and introduced into clinical practice as a therapeutic agent two years later [2]. Chloramphenicol was notably efficient against 95% of gram-negative bacteria and against a variety of gram-positive aerobic strains The extensive use of chloramphenicol in the treatment of infectious diseases in humans and animals has caused increasing concerns about its effect on human health. Its bitter taste has led to the development of chloramphenicol alternatives. Many studies have addressed the derivatization of chloramphenicol, including selective esterification of the hydroxyl

Methods

Results

Conclusion