Publication Date


Document Type


First Advisor

Horn, James R.

Degree Name

M.S. (Master of Science)

Legacy Department

Department of Chemistry and Biochemistry


VHH domains are the antigen-binding units found in heavy-chain-only antibodies from the Camelidae family (e.g., camels and llamas). They are considered a unique minimalist antibody scaffold due to the lack of a variable light chain domain. Despite having half the antigen-binding interface of conventional antibodies, these VHH antibodies bind their target antigens with affinities comparable to conventional antibodies. This project explored how flexible linkers connecting an anti-GFP VHH (αGFP VHH) domain to its antigen (EGFP) enhanced antigen stability and influenced oligomeric assembly. It was hypothesized that linking the anti-GFP VHH antibody to its target antigen (EGFP) will increase the local concentration of antibody, thus increasing the stability of the antigen. Consequently, we wanted to test the effect of linker length on both thermal/chemical stability and oligomeric state. The results confirm that a flexible linker between the EGFP antigen and the anti-GFP VHH stabilized EGFP with an observed increase in melting temperature of 8-11°C. In addition to showing enhanced stability, the linker length affected the oligomeric assemblies of the EGFP-αGFP VHH fusions and induced the formation of monomers and higher-order oligomers EGFP. Since proteins are used in a wide range of applications, including medical imaging, therapeutics, and diagnostics, VHH fusions may serve as useful modules to extend the function of proteins across a wider range the environmental conditions (e.g., high temperature and harsh chemical conditions).


117 pages




Northern Illinois University

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