Riboflavin Interference in Monitoring Protein Production in Pichia pastoris
Author Information
Author(s): Surribas Anna, Resina David, Ferrer Pau, Valero Francisco
Primary Institution: Departament d'Enginyeria Química, Universitat Autònoma de Barcelona
Hypothesis
Can riboflavin interfere with the monitoring of GFP fusion proteins in Pichia pastoris?
Conclusion
Riboflavin significantly interferes with the GFP fluorescence signal, complicating the monitoring of extracellular protein production in Pichia pastoris.
Supporting Evidence
- The GFP-ROL fusion protein was successfully produced in Pichia pastoris.
- Riboflavin was identified as a major interfering component with the GFP signal.
- Ultrafiltration was necessary to remove riboflavin for accurate GFP fluorescence measurement.
- Extracellular GFP fluorescence correlated with lipase activity after riboflavin removal.
Takeaway
This study found that riboflavin can mess up the signals we use to see how much protein is being made in yeast, which is important for scientists trying to measure protein production.
Methodology
The study involved producing a GFP-ROL fusion protein in Pichia pastoris and using multiwavelength spectrofluorometry to monitor its production.
Limitations
The study's findings may not be applicable to all GFP variants or other systems, and ultrafiltration may not be practical for on-line monitoring.
Digital Object Identifier (DOI)
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