Determining the properties of protein folding intermediates is critical to understanding how proteins fold and why they misfold, and to identifying the determinants of protein aggregation. Misfolding or off-pathway folding events, particularly those leading to aggregation, have been identified as the causative agents in several human diseases including cystic fibrosis, Alzheimer's disease, and prion spongiform encephalopathies such as Creutzfeldt-Jacob disease. A better understanding of the interactions which promote folding and prevent aggregation will enhance Dr. Robinson's ability to design inhibitors and therapeutics for aggregation-driven diseases. Despite increasing progress in identifying features of intermediates of small well-behaved proteins, understanding folding and assembly of large multi-domain proteins is still a critical problem, particularly in the post-genomic area. There is limited ability to translate the knowledge and techniques gained in small protein systems to those of large, complex proteins and oligomers. Structural and functional genomics initiatives will require a better understanding of the folding and assembly of oligomers and other complex protein systems. It is likely that such proteins require novel mechanisms which assist folding, and novel techniques to evaluate their intermediates. Dr. Robinson is investigating assembly intermediates of P22 tailspike protein, and a novel class of interactions between cysteine residues that direct folding and assembly through transient intersubunit disulfide bond formation. P22 tailspike protein is an homotrimer of 666 amino acids per monomer chain. Tailspike forms transient disulfide bonds in an intermediate on the folding and assembly pathway in vivo and in vitro. Disulfide bond formations appear to facilitate folding and disfavor aggregation, and may help in beta-sheet alignment during oligomerization. In order to understand the role of transient disulfide bond formation in tailspike folding and assembly, Dr. Robinson's research program will identify amino acid determinants of disulfide bond formation in vivo and in vitro. Dr. Robinson will also develop a novel method which combines cyanylation, proteolytic cleavage, and MALDI mass spectroscopy to map disulfide bond connectivity in tailspike assembly intermediates.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Molecular and Cellular Biophysics Study Section (BBCA)
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Wehrle, Janna P
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University of Delaware
Engineering (All Types)
Schools of Engineering
United States
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Kim, Junghwa; Robinson, Anne Skaja (2006) Dissociation of intermolecular disulfide bonds in P22 tailspike protein intermediates in the presence of SDS. Protein Sci 15:1791-3
Gage, Matthew J; Zak, Jennifer L; Robinson, Anne Skaja (2005) Three amino acids that are critical to formation and stability of the P22 tailspike trimer. Protein Sci 14:2333-43
Danek, B L; Robinson, A S (2004) P22 tailspike trimer assembly is governed by interchain redox associations. Biochim Biophys Acta 1700:105-16
Danek, Brenda L; Robinson, Anne Skaja (2003) Nonnative interactions between cysteines direct productive assembly of P22 tailspike protein. Biophys J 85:3237-47
Gage, Matthew J; Robinson, Anne Skaja (2003) C-terminal hydrophobic interactions play a critical role in oligomeric assembly of the P22 tailspike trimer. Protein Sci 12:2732-47