How to Use Asp-N or Glu-C Alone or Combined for Protein Digestion? Dosage vs. Trypsin’s 30:1 Ratio
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Dosage: A protein-to-enzyme ratio of 50:1 is recommended.
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Digestion Conditions: Incubate at approximately pH 8.0 and 37°C, typically overnight.
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Dosage: A protein-to-enzyme ratio in the range of 20:1 to 30:1 is recommended.
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Digestion Conditions: Optimal at pH 7.8, incubated at 37°C overnight.
The following guidelines outline recommended protocols for the usage and dosage of Asp-N and Glu-C in protein digestion experiments:
Using Asp-N or Glu-C Alone
1. Asp-N
2. Glu-C
Sequential Digestion with Asp-N and Glu-C
1. Perform an initial digestion using Asp-N under the conditions described above (50:1 ratio, pH 8.0, 37°C, overnight).
2. Following the initial step, add Glu-C and continue the digestion overnight, using a protein-to-enzyme ratio between 20:1 and 30:1.
This sequential strategy enables broader coverage of potential proteolytic cleavage sites, thereby enhancing both sequence coverage and digestion efficiency. These parameters can be further optimized based on specific experimental objectives and enzyme characteristics.
MtoZ Biolabs, an integrated chromatography and mass spectrometry (MS) services provider.
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