Dear Owais Ahmad

This is a tough one to answer. It depends on numerous factors:

-What is small? A bit like the discussion where peptides end, and proteins start.

-Type of peptide/protein. What is the secondary structure, is there some kind of domain or domains formation, etc.

-Is the folding dependent on other factors yes or no. Is there a chaperone, does it require binding/interaction with lipids, is there a monomeric or oligomeric formation, etc.

A good starting point is:

Jackson, S. E. (1998). How do small single-domain proteins fold?. Folding and Design, 3(4), R81-R91. https://www.cell.com/foldes/pdf/S1359-0278(98)00033-9.pdf and

Dobson, C. M. (2004, February). Principles of protein folding, misfolding and aggregation. In Seminars in cell & developmental biology (Vol. 15, No. 1, pp. 3-16). Academic Press. https://home.cc.umanitoba.ca/~joneil/Folding-2004.pdf

All I know is that smaller proteins or better peptides are obtained by:

-Chemical synthesis

-Isolation from a source (a bacterium, from poisons like venom etc.)

-By fragmentation of a bigger protein

See for more info for example:

Akbarian, M., Khani, A., Eghbalpour, S., & Uversky, V. N. (2022). Bioactive peptides: Synthesis, sources, applications, and proposed mechanisms of action. International journal of molecular sciences, 23(3), 1445. https://doi.org/10.3390/ijms23031445

I don’t have a direct example of a one-to-one comparison of a synthetic peptide and the same peptide from a natural source on the top of my mind, but I do know numerous examples of peptides that are obtained by chemical synthesis that exhibit some kind of bioactivity, see for examples:

Sánchez, A., & Vázquez, A. (2017). Bioactive peptides: A review. Food quality and safety, 1(1), 29-46. Article Bioactive peptides: A review

Perhaps someone else here on RG recalls an example of a direct comparison.

Best regards.

Hi Owais Ahmad

If you are working with small proteins, then the answer is generally yes. If bioactivity of a protein is dependent on its fold, we can assume that biologically active synthetic proteins must have folded properly.

Many synthetic peptides (short) were tested in animal models and they work fine. Larger proteins cannot be synthesized chemically, so some groups made semi-synthetic proteins like the ones mentioned below. All such proteins have worked well in vivo. In some papers, you may find CD spectra for such semi-synthetic proteins comparing them with native or pharmaceutical grade proteins; they usually acquire native folds.

Examples:

1) Inteins - a fragment of the protein is synthesized chemically and linked with remaining portion of protein which is expressed recombinantly. (PMIDs: 12797783, 32424098)

2) Bioconjugation - some proteins are chemically modified with unnatural groups and then purified on reverse phase HPLC with organic solvents containing TFA. When reconstituted in buffer, they show activity in mice/rats. (My work, PMIDs: 29192764, 37369330)

Dear Rob Keller ,

Thank you for your response. I inquired about this matter because my current laboratory lacks the necessary infrastructure and chemical reagents for gene cloning and purification of translated products. My peptides and proteins consist of fewer than 100 amino acids. I was concerned about the correct folding of the synthesized proteins/peptides, and your insights will assist me in making an informed decision.

Dear Karthik Nadendla ,

Thank you for your answer. Based on your guidance, I can confidently conclude that smaller proteins and peptides are likely to fold correctly in solution.