Progress toward purification of the peptide

We were able to determine that we have produced pure peptide from cell extracts by LC/MS. That is a liquid chromatograph/mass spectrometer. We used some of the chemically synthesized peptide to determine a method for the LC/MS analysis. Once the method was determined, we tested some of our processed cell extracts. The method involved centrifuging the cells from the fermentation broth, breaking open the cells and separating the cell debris from the liquid in the cells, or cytoplasm. The gel image shows that (lane 1 is the molecular weight marker, lane 4 is when the cells were given a chemical to cause the expression of our peptide chain and lane 8 shows the dark band of our expressed peptide chain that was harvested at the end of the fermentation.

 

From the centrifuged cell pellets we were able to separate out the peptide chain and then cleave it apart with an enzyme. Only 15 minutes incubation with the enzyme showed that we were obtaining pure peptide by LC/MS. This week we are concentrating the peptide and will send it for activity analysis in Texas.