Hello everyone,
I want to isolate antimicrobial peptides from plants but, I don't have a protease inhibitor cocktail right now and possibly not in the near future.
Is there any alternative method to isolate antimicrobial peptides from plants without using protease inhibitor cocktail ? what can I use for isolation of antimicrobial peptides instead of using protease inhibitor cocktail ?
Thank you in advance
Hi,
If you extract and purify the peptides at low temperature, you might be able to suppress the protease activity enough to get a good yield.
You could start by grinding the plant matter under liquid nitrogen, which would prevent any proteolysis by keeping everything frozen. Once the liquid nitrogen evaporated, you could extract the powder. Since antimicrobial peptides are relatively small compared to proteins, you probably don't have to be concerned about retaining their three-dimensional structure. Therefore, you could extract them under conditions that would destroy the activity of proteolytic enzymes, such as by using an organic solvent such as alcohol or acetone, which could be evaporated easily later.
After extraction of the peptides using predefined buffers without denaturants (if native peptide structures are needed) you may promptly load the extract to C18 resin with low pore sizes (approx.90-110 A max). Probably flow through fraction will contain the proteins (including proteases) which cannot diffuse into the pores of the resin due to their radius and size exclusion effect. Look for the details by typing flash chromatography peptide purification...
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