FAQ: Why is my protein degraded? When I denature and add SDS all I see on my SDS-PAGE is a smear or no protein. Can a protease inhibitor cocktail be used in a PNGase A reaction?

When a protein is denatured it is more susceptible to cleavage by proteases. For this reason a protease cocktail containing the following can be used in a PNGase A reaction protocol:

Aprotinin (10 μg/ml final concentration)
Benzamidine (1mM final concentration)
Pepstatin (10 μg/ml final concentration)
Leupeptin (1μM final concentration)
EGTA (1 mM final concentration)
EDTA (1 mM final concentration)
PMSF (1 mM final concentration)

Make a 1000X concentrated stock of each inhibitor in water; except Pepstatin which should be dissolved in methanol.

Note: PMSF has the ability to modify basic residues on glycoprotein substrates.

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Related Products:
PNGase A