Abstract
The purification and characterization of PGM (Phosphoglucomutase) from Cordyceps militaris (C. militaris) was investigated. PGM was purified using a combination of ultrafiltration, salting-out and ion exchange chromatography resulting in 4.23-fold enhancement of activity with a recovery of 20.01%. Molecular mass was 50.01 kDa by SDS-PAGE. The optimal activity was achieved at pH 7.5 and 30 °C with NADPH as substrate. The results showed that SDS, DTT Li+, Cu2+, Na+, Mn2+ and Al3+ were effective PGM inhibitors; whereas glycerol, Zn2+, Mg2+, Ca2+, Fe2+ and Fe3+ could enhance the activity of PGM, and the Km and Vmax values were 11.62 mmol/L and 416.67 U/mL, respectively. At the same time, qRT-PCR was used to test the changes of mRNA transcription level of PGM gene encoding under two fermentation conditions: basic medium and optimized medium. The relative quantitative results of PGM target genes resulting in 2.60-fold enhancement than the control group.
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This work was financially supported by the National Natural Science Foundation of China (31871791), the key program of the Natural Science Foundation of Tianjin (16JCZDJC34100), Technology Program of Tianjin, China, (18ZYPTJC00020) and the key program of the Foundation of Tianjin Educational Committee (2018ZD06).
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ZZY conceived and designed research. JYP conducted experiments. LCY contributed new reagents or analytical tools. JYP and GXQ analyzed data. GXQ wrote the manuscript. All authors read and approved the manuscript.
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Geng, XQ., Ji, YP., Liu, CY. et al. Enzymatic characterization and validation of gene expression of phosphoglucomutase from Cordyceps militaris. Biotechnol Lett 43, 177–192 (2021). https://doi.org/10.1007/s10529-020-02981-3
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DOI: https://doi.org/10.1007/s10529-020-02981-3