I am doing my research on isolation of yeast from fermented food to extract antimicrobial yeast peptides. I need standardized protocol for the extraction of protein from yeast strains.
There are a few different methods for extracting proteins from yeast strains, and the specific protocol will depend on the type of yeast and the intended use of the proteins.
Here is a general protocol for extracting proteins from yeast:
Start by growing the yeast strains in liquid culture or agar plates.
Harvest the yeast cells by centrifugation at 3000-5000 x g for 10-15 minutes.
Resuspend the cell pellet in a buffer such as Tris-HCL or phosphate-buffered saline (PBS) and lyse the cells by mechanical means such as French press, homogenizer, bead-beater, or ultrasonicator.
Clarify the lysate by centrifugation at 10,000-15,000 x g for 20-30 minutes.
The supernatant contains the soluble proteins. To obtain the membrane proteins, pellet the yeast cells by centrifugation and resuspend the cells in a buffer containing a detergent such as Triton-X100 or SDS. Sonicate the cells to disrupt them and centrifuge the lysate to separate the membrane proteins
The proteins can be further purified by various techniques such as centrifugation, precipitation, chromatography, or electrophoresis.
To purify the antimicrobial peptides, you can use techniques such as reverse-phase HPLC, size-exclusion chromatography, or ion-exchange chromatography.
Finally, concentrate and desalt the purified proteins using techniques such as ultrafiltration or dialysis.
It's important to keep in mind that proteins can be denatured, degraded or lose activity during the extraction process, so it's important to optimize the protocol for your specific yeast strains and proteins of interest. Additionally, It's worth noting that the best extraction method should be chosen based on the type of yeast cell wall and the type of antimicrobial peptides you're trying to extract.