Enzymatic characterization and validation of gene expression of phosphoglucomutase from Cordyceps militaris

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 K_m and V_max 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.