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Production, Purification, and Characterization of Polygalacturonase from Rhizomucor pusillus Isolated from
Mohd Asif Siddiqui1, Veena Pande, Mohammad Arif
1Department of Biotechnology, Kumaun University, Campus Bhimtal, Nainital 263136, India.
Abstract:
A thermophilic fungal strain producing polygalacturonase was isolated after primary screening of 40 different isolates. The fungus was identified as Rhizomucor pusilis by Microbial Type Culture Collection (MTCC), Chandigarh, India. An extracellular polygalacturonase (PGase) from R. pusilis was purified to homogeneity by two chromatographic steps using Sephadex G-200 and Sephacryl S-100. The purified enzyme was a monomer with a molecular weight of 32 kDa. The PGase was optimally active at 55°C and at pH 5.0. It was stable up to 50°C for 120 min of incubation and pH condition between 4.0 and 5.0. The stability of PGase decreases rapidly above 60°C and above pH 5.0. The apparent K(m) and V(max) values were 0.22 mg/mL and 4.34 U/mL, respectively. It was the first time that a polygalacturonase enzyme was purified in this species. It would be worthwhile to exploit this strain for polygalacturonase production. Polygalacturonase from this strain can be recommended for the commercial production because of its constitutive and less catabolically repressive nature, thermostability, wide range of pH, and lower K(m) properties. However, scale-up studies are needed for the better output for commercial production.
Insights
A thermophilic fungus, Rhizomucor pusilis, was identified to produce polygalacturonase (PGase). This PGase enzyme exhibits optimal activity and stability at high temperatures and a specific pH range, making it suitable for commercial applications.
Area of Science:
- Enzymology
- Microbiology
- Biotechnology
Background:
- Polygalacturonase (PGase) enzymes are crucial in pectin degradation.
- Thermophilic fungi are a promising source of robust industrial enzymes.
- Limited research exists on PGase from Rhizomucor pusilis.
Purpose of the Study:
- To isolate and identify a thermophilic fungal strain producing polygalacturonase.
- To purify and characterize the extracellular polygalacturonase (PGase) from Rhizomucor pusilis.
- To evaluate the potential of this PGase for commercial production.
Main Methods:
- Screening of 40 fungal isolates for polygalacturonase activity.
- Identification of the fungal strain using Microbial Type Culture Collection (MTCC).
- Purification of extracellular PGase via Sephadex G-200 and Sephacryl S-100 chromatography.
- Enzyme characterization including molecular weight, optimal activity, stability, K(m), and V(max) determination.
Main Results:
- A thermophilic fungal strain, identified as Rhizomucor pusilis, was isolated.
- Extracellular PGase was purified to homogeneity with a molecular weight of 32 kDa.
- Optimal activity was observed at 55°C and pH 5.0; enzyme stability was noted between pH 4.0-5.0 and up to 50°C for 120 min.
- Apparent K(m) and V(max) were 0.22 mg/mL and 4.34 U/mL, respectively.
- This is the first reported purification of polygalacturonase from this species.
Conclusions:
- Rhizomucor pusilis produces a thermostable polygalacturonase suitable for industrial applications.
- The enzyme's properties (thermostability, pH range, low K(m)) recommend it for commercial production.
- Further scale-up studies are necessary to optimize output for commercial viability.
