Plant-Derived Hydrolysates Are a Suitable Replacement for Tryptone N1 in Recombinant Protein Expression Using Human...

Area of Science:

• Biotechnology
• Cell Biology
• Biochemistry

Background:

• Human embryonic kidney (HEK293) cells are widely used for recombinant protein expression.
• Tryptone N1 (TN1), an animal-derived casein hydrolysate, is commonly added post-transfection to optimize yields.
• Regulatory challenges with animal by-products necessitate exploring plant-based alternatives.

Purpose of the Study:

• To evaluate plant-derived hydrolysates as potential replacements for TN1 in HEK293 cell cultures.
• To compare the impact of plant-based vs. animal-based hydrolysates on cell viability and recombinant protein expression.

Main Methods:

• HEK293-6E cells were transfected using polyethyleneimine (PEI) in suspension, serum-free medium.
• Cultures were supplemented with pea hydrolysate, broad bean hydrolysate, TN1, or no hydrolysate.
• Cell counts, viability, and His-tagged recombinant protein expression were quantified via stain-free Western blotting.

Main Results:

• Plant-derived hydrolysates and TN1 maintained similar cell viability compared to the no-hydrolysate control.
• Pea and broad bean hydrolysates yielded comparable levels of recombinant protein expression to TN1.
• All tested hydrolysates demonstrated superior performance over the no-hydrolysate control.

Conclusions:

• Plant-derived hydrolysates are a viable and sustainable alternative to animal-derived TN1 for recombinant protein production.
• These findings support the use of plant-based peptides to overcome regulatory hurdles associated with animal by-products.