Related Experiment Videos
Synthesis of mumps virus nucleocapsid protein in yeast Pichia pastoris
Rimantas Slibinskas1, Aurelija Zvirbliene, Alma Gedvilaite
1Institute of Biotechnology, V Graiciuno 8, Vilnius LT-2028, Lithuania.
Abstract:
The expression of mumps virus nucleocapsid protein in yeast Pichia pastoris was investigated. Viral nucleocapsid proteins usually elicit a strong long-term humoral immune response in patients and experimental animals. Therefore, the detection of antibodies specific to mumps virus nucleoprotein can play an important role in immunoassays for mumps diagnosis. For producing a high-level of recombinant mumps virus nucleoprotein the expression system of yeast P. pastoris was employed. The recombinant nucleocapsid protein was purified by cesium chloride ultracentrifugation of yeast lysates. Electron microscopy of the purified recombinant nucleocapsid protein revealed a herring-bone structure similar to the one discovered in mammalian cells infected with mumps virus. The yield of purified nucleocapsid-like particles from P. pastoris constituted 2.1 mg per 1 g of wet biomass and was considerably higher in comparison to the other expression systems.
Insights
Researchers expressed mumps virus nucleocapsid protein in Pichia pastoris yeast. This high-yield system produced purified nucleocapsid-like particles, valuable for mumps diagnostic immunoassays.
Area of Science:
- Virology
- Molecular Biology
- Biotechnology
Background:
- Mumps virus nucleocapsid proteins induce strong humoral immune responses.
- Antibodies to mumps nucleoprotein are crucial for diagnostic immunoassays.
- Efficient production of recombinant nucleocapsid protein is needed.
Purpose of the Study:
- To investigate the expression of mumps virus nucleocapsid protein in the yeast Pichia pastoris.
- To develop a high-level production system for recombinant mumps virus nucleoprotein.
- To characterize the purified recombinant nucleocapsid protein.
Main Methods:
- Utilized the Pichia pastoris expression system for recombinant protein production.
- Purified the recombinant nucleocapsid protein using cesium chloride ultracentrifugation.
- Analyzed the structure of purified particles via electron microscopy.
Main Results:
- Achieved a high yield of purified nucleocapsid-like particles (2.1 mg/g wet biomass).
- Electron microscopy confirmed a herring-bone structure, similar to native mumps virus.
- The P. pastoris system demonstrated superior yield compared to other expression systems.
Conclusions:
- Pichia pastoris is an effective host for high-level expression of mumps virus nucleocapsid protein.
- The produced recombinant protein possesses structural integrity relevant for diagnostic applications.
- This system offers a promising platform for producing reagents for mumps diagnosis.