Open Collections

The study of homo- and hetero-substituted de novo four-helix bundle proteins

University of British Columbia

The ability to design, synthesize and characterize de novo proteins can help facilitate the understanding of how individual amino acids contribute to the stability and structure of a protein. The de novo approach can be extended to include the use of templates, which assist in the organization of the peptides to form predetermined three-dimensional structures. These template assembled de novo proteins have been named "caviteins" (cavitand + protein).[See thesis for diagram]. One of the challenges in this area of research is the ability to design and synthesize native-like de novo proteins. Previously two caviteins, LG2 and LG3, had shown some nativelike characteristics, although the evidence for a completely native-like structure remained debatable. The approach to identify a native-like structure was to design a corresponding protein that would exhibit non native-like properties. The leucine residues of LG2 and LG3 were replaced with norleucine residues in NG2 and NG3, respectively. The norleucine-based caviteins were less native-like in structure, as speculated, than their leucine-based counterparts. In the past, the designed caviteins were limited to having only one type of peptide sequence attached within one bundle. Here, the design of a hetero-TASP, i.e. two different sequences within one bundle, was explored, and provided a means to create various de novo proteins, including an anti-parallel four-helix bundle. The hetero-TASPs were characterized and found to exhibit different native-like properties depending on the attached peptide sequences, and helix orientations. Lastly, the N - and C-capping efficiency of glycine was examined. Caviteins having, peptides linked to the cavitand template via their N - and C-termini, and with and without glycine caps were synthesized and characterized. It was found that the caviteins lacking glycine caps at their respective helix termini were comparable in stability with their capped-counterparts, which was contrary to what was hypothesized. It has been shown that subtle changes in the peptide sequence, linker and helix orientation have dramatic effects on the overall cavitein structure and stability. Since many of the factors underlying the stability and structure of four-helix bundles are now well understood, it would be exciting to undertake the challenges of designing caviteins with specific applications.

Text

2011-01-27

For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use.

http://hdl.handle.net/2429/30901

Chemistry

University of British Columbia

Graduate