Microsequence analysis of peptides and proteins. VI. A continuous flow reactor for sample concentration and sequence...

Area of Science:

• Biochemistry
• Analytical Chemistry
• Chemical Engineering

Background:

• Conventional microsequence analysis relies on cartridge/glass fiber disk designs.
• Existing methods can suffer from high background noise and require precycling of reagents.
• Handling low picomole amounts of peptides and proteins presents analytical challenges.

Purpose of the Study:

• To design and test a novel continuous flow reactor (CFR) for enhanced microsequence analysis.
• To improve upon existing methods by reducing background noise and increasing yields.
• To facilitate sample handling and analysis of low picomole quantities of peptides and proteins.

Main Methods:

• A continuous flow reactor (CFR) was constructed using Teflon tubing packed with Polybrene-coated silica.
• The CFR design allows for immobilization of peptide/protein substrates and automated Edman chemistry.
• Reverse-phase packing was incorporated for direct sample concentration onto the CFR.

Main Results:

• The CFR demonstrated significantly lower background noise, particularly reducing the diphenylthiourea peak.
• Higher initial and repetitive yields were achieved compared to conventional cartridge/glass fiber disk designs.
• Direct concentration of 1.0-ml samples (200 pmol) onto the CFR yielded 95-100% recovery.

Conclusions:

• The developed continuous flow reactor offers a cost-effective and disposable solution for microsequence analysis.
• The CFR design enhances sample handling and improves the accuracy of peptide and protein sequencing.
• This methodology shows promise for the analysis of trace amounts of peptides and proteins.