Beyond Antibodies: Development of a Novel Protein Scaffold Based on Human Chaperonin 10

• Published in: Scientific reports Vol. 6; no. 1; p. 37348
• Main Authors: Alsultan, Abdulkarim M., Chin, David Y., Howard, Christopher B., de Bakker, Christopher J., Jones, Martina L., Mahler, Stephen M.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 22.11.2016; Nature Publishing Group
• Subjects: 13; 13/106; 13/31; 14; 631/45/612/1981; 631/61/338/469; 631/61/51/2314; 82/80; 82/83; Affinity; Antibodies; Avidity; Binding Sites; Blood Coagulation - drug effects; Cancer; CD44 antigen; Chaperonin 10 - chemistry; Clotting; Coagulation factor VIIa; Coagulation factors; E coli; Factor VIIa - pharmacology; Hemorrhage; Humanities and Social Sciences; Humans; Hyaluronan Receptors - metabolism; Models, Molecular; Molecular Dynamics Simulation; multidisciplinary; Peptide Library; Peptides; Peptides - chemistry; Peptides - pharmacology; Pregnancy Proteins - chemistry; Protein Binding; Protein Structure, Quaternary; Proteins; Prothrombin; Science; Suppressor Factors, Immunologic - chemistry
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep37348

Human Chaperonin 10 (hCpn10) was utilised as a novel scaffold for presenting peptides of therapeutic and diagnostic significance. Molecular dynamic simulations and protein sizing analyses identified a peptide linker (P1) optimal for the formation of the quarternary hCpn10 heptamer structure. hCpn10 scaffold displaying peptides targeting Factor VIIa (CE76 -P1 ) and CD44 (CP7) were expressed in E. coli . Functional studies of CE76 -P1 indicated nanomolar affinity for Factor VIIa (3 nM) similar to the E-76 peptide (6 nM), with undetectable binding to Factor X. CE76 -P1 was a potent inhibitor of FX activity (via inhibition of Factor VIIa) and prolonged clot formation 4 times longer than achieved by E-76 peptide as determined by prothrombin time (PT) assays. This improvement in clotting function by CE76 -P1 , highlights the advantages of a heptamer-based scaffold for improving avidity by multiple peptide presentation. In another example of hCPn10 utility as a scaffold, CP7 bound to native CD44 overexpressed on cancer cells and bound rCD44 with high affinity (K D 9.6 nM). The ability to present various peptides through substitution of the hCpn10 mobile loop demonstrates its utility as a novel protein scaffold.