原文链接：Characterization of a novel esterase and construction of a Rhodococcus-Burkholderia consortium capable of catabolism bis (2-hydroxyethyl) terephthalate - ScienceDirect

亮点

• 鉴定了BHET降解菌Rhodococcus biphenylivorans GA1和Burkholderia sp. EG1。。

• 克隆了一种新的酯酶基因betH。

• BetH对BHET具有很高的催化活性。

• Rhodococcus-Burkholderia菌群通过互惠互利降解高浓度的BHET。

摘要

双(2-羟乙基)对苯二甲酸酯（BHET）是通过对聚对苯二甲酸乙二醇酯（PET）进行酶解或化学降解而产生的主要化合物之一。然而，对BHET微生物代谢的认识不足是限制PET生物回收利用的主要因素之一。本研究中，从Rhodococcus biphenylivorans GA1和Burkholderia sp. EG1中分离并鉴定出了BHET降解菌株，它们可以利用BHET作为唯一的碳源生长。此外，从GA1菌株中克隆了一种新型酯酶基因betH，它编码一种在30°C和pH 7.0下具有最高活性的BHET水解酯酶。此外，含有GA1菌株和EG1菌株的共培养体系可以完全降解高浓度的BHET，消除了由于中间代谢产物乙二醇（EG）积累而引起的对GA1菌株的抑制。本研究为PET生物回收利用提供了潜在菌株和可行的策略。

Fig. 1. Characteristics of BHET degradation by strain GA1. (A) Transparent halo formed by strain GA1 on LB plate with 1.2 mM BHET. (B, C) The effect of temperature and pH on the degradation of BHET by strain GA1. (D) Utilization of BHET or TPA as the sole carbon source for growth by strain GA1. The initial concentration of BHET and TPA was 1.2 mM, and 1.2 mM substrates were added respectively after incubation 21h. (E) Effects of addition 1.2 mM EG on the growth and BHET degradation of strain GA1.

Fig. 2. Metabolic pathway analysis of degradation of BHET by strain GA1. (A) HPLC detection results. (B–D) The peaks of compound I (BHET), compound II (MHET) and compound Ⅲ (TPA) were further analyzed by MS/MS.

Fig. 3. Analysis of metabolic pathways based on the genome of strain GA1. (A) Genomic context of BHET catabolism-related genes in strain GA1. (B) Proposed BHET-catabolic pathway in strain GA1.

Fig. 4. Cloning of BHET hydrolase gene and study on BetH enzymatic characteristics. (A) Transparent halos formed by tested strains or crude enzyme on LB agar supplemented with 1.2 mM BHET. (B) Phylogenetic comparison of BetH with selected esterase/lipase. (C) SDS-PAGE analysis of the purified BetH. Effects of temperature (D) and pH (E) on enzyme activity of BetH. (F) The kinetic curve of BetH with BHET as substrate. Error bars represent the standard errors for three replicates.

Fig. 5. (A) Utilization of EG as the sole carbon source for growth by strain EG1. (B) Neighbour-joining tree based on 16S rRNA gene sequences, showing the relationship between strain EG1 and related taxa.

Fig. 6. Complete degradation of BHET by a co-culture system. (A) Schematic representation of the BETH catabolism mechanisms by Rhodococcus biphenylivorans and Burkholderia sp. co-culture. (B) Utilization of BHET as the sole carbon source for growth by Rhodococcus biphenylivorans and Burkholderia sp. co-culture.