Mining of key genes for xylose metabolism and cloning, expression, and enzymatic characterization of XylA in Bacillus coagulans.
- Author:
Yiwen ZHANG
1
;
Yajie ZHANG
1
;
Manxin CHEN
1
;
Xiaojun GUO
2
;
Baocheng ZHU
2
;
Yuming ZHANG
1
Author Information
- Publication Type:Journal Article
- Keywords: Bacillus coagulans; biorefinery; lactic acid; transcriptomic analysis; xylose isomerase
- MeSH: Xylose/metabolism*; Cloning, Molecular; Bacillus coagulans/enzymology*; Aldose-Ketose Isomerases/metabolism*; Fermentation; Bacterial Proteins/metabolism*; Glucose/metabolism*
- From: Chinese Journal of Biotechnology 2025;41(10):3876-3890
- CountryChina
- Language:Chinese
- Abstract: Bacillus coagulans can utilize the hydrolyzed carbon source of agricultural waste to produce lactic acid via a homofermentative pathway. However, a significant carbon source metabolic repression effect was observed when the strain metabolized mixed sugars (glucose and xylose), reducing the productivity of lactic acid. In this study, we obtained the fermentation conditions for the simultaneous utilization of the mixed sugars by B. coagulans by changing the ratio of glucose to xylose in the medium. Through transcriptome sequencing, several key genes responsible for xylose utilization were identified. The critical role of xylose isomerase (XylA, EC 5.3.1.5) in the synchronous utilization of glucose/xylose in B. coagulans was investigated via qRT-PCR (quantitative real-time polymerase chain reaction). Subsequently, the heterologous expression and characterization of the XylA-encoding gene (XylA) were conducted. It was determined that the gene encoded a protein composed of 440 amino acid residues. The secondary structure of the encoded protein was predominantly composed of α-helixes and random coils, while the higher structure of the protein was identified as a homotetramer. Then, XylA was cloned and expressed in Escherichia coli BL21(DE3), and the recombinant protein Bc-XlyA was obtained with a molecular weight of approximately 50 kDa. The optimal pH and temperature of Bc-XylA were 8.0 and 60 ℃, respectively, and Mn2+, Mg2+, and Co2+ had positive effects on the activity of Bc-XlyA. The present study provides scientific data on the molecular modification of B. coagulans, offering theoretical support for the efficient utilization of xylose in the strain.
