Implementation of a Practical Teaching Course on Protein Engineering.

Area of Science:

• Biotechnology
• Molecular Biology
• Biochemistry

Background:

• Protein engineering is crucial for developing proteins for diverse applications.
• Recombinant protein production in Escherichia coli is a common method.
• Practical laboratory courses are essential for training in protein purification techniques.

Purpose of the Study:

• To present a practical teaching course on recombinant protein production.
• To demonstrate fundamental techniques for expressing, purifying, and analyzing enhanced green fluorescent protein (eGFP).
• To enable students to gain hands-on experience in protein engineering.

Main Methods:

• Sequential laboratory sessions covering bacterial growth, cell lysis (sonication), and eGFP purification (affinity chromatography, dialysis).
• Protein quantification using bicinchoninic acid (BCA) and fluorometry assays.
• Qualitative analysis via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).

Main Results:

• Students successfully isolated eGFP from cell lysate with purity levels up to 72%.
• Mass balance analysis indicated eGFP yields of up to 46% were achieved.
• Sensitivity analysis identified critical steps in the downstream processing.

Conclusions:

• The practical course effectively trained students in essential protein engineering techniques.
• The established methodologies are suitable for producing and analyzing recombinant eGFP.
• Understanding downstream processing is key to optimizing protein purification yields.