Computational design of orthogonal antiparallel homodimeric coiled coils

Living cells integrate a vast array of protein-protein interactions (PPIs) to govern cellular functions. For instance, PPIs are critical to biosynthesis, nanostructural assembly, and in processing environmental stimuli through cell-signaling pathways. As fields such as synthetic biology and protein engineering mature they seek to mimic and expand the functions found in living systems that integrate PPIs. A critical feature to many PPIs that are integrated together to perform a complex function is orthogonality, i.e. PPIs that do not cross interact with each other. The engineering of orthogonal PPIs is thus an alluring problem. Since it not only tests our understanding of molecular specificity by having to stabilize and destabilize interactions simultaneously. The results of the design process can also have interesting applications in synthetic biology or bionanotechnology. The coiled coil, a rope-like structure made of helices, is a PPI ubiquitously found in biological systems and is an attractive fold for engineering orthogonal PPIs. Though the coiled coil is well studied, destabilization of undesired interactions still remains challenging. In this thesis I will discuss strategies for obtaining orthogonal PPIs, and describe the current sequence-to-structure relationships known about coiled coils. I will then introduce the computational multistate design framework, CLASSY, and explain how I applied it to the computational design of six orthogonal antiparallel homodimeric coiled coils. Five of these designed sequences were experimentally tested, of which only three of the sequences adopted the target antiparallel homodimer topology. All three of these sequences, as well as a previously designed antiparallel homodimer, were tested for cross reactivity in a pairwise manner. None of these sequences appeared to cross react. The sequences that failed to adopt the antiparallel topology highlight the need for improving our computational design framework. In the final chapter I will discuss strategies to improve our models, and applications for orthogonal antiparallel coiled coils.

Department dc:contributor.department

Massachusetts Institute of Technology. Computational and Systems Biology Program.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2014

Author dc:creator

• Negron, Christopher

Advisor dc:contributor.advisor

• Amy E. Keating.

• Computational and Systems Biology Program.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/93805

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/93805