Combinatorial, rational, and bioinformatics approaches to engineering cystine-rich proteins
- Proteins are the most structurally and functionally diverse class of biological molecules. By exploiting this diversity, nature has evolved proteins for complex biological functions such as molecular recognition and catalysis. The features of proteins that contribute to their structure, stability, and function have been identified through extensive experimental work. However, the principles governing the precise relationships between a protein's sequence, structure, and function remain largely unknown. Here we discuss the application of combinatorial, rational, and bioinformatics techniques for engineering two model cystine-rich proteins: Ecballium elaterium trypsin inhibitor-II (EETI) knottin and epidermal growth factor (EGF). With this multi-faceted engineering approach, we have assessed the sequence plasticity of the structure of EETI and have predicted and experimentally validated a set of sequence criteria that facilitate proper folding of the knottin structure. Using a similar array of protein engineering techniques we have developed EGF variants with modulated receptor binding properties. These EGF variants will serve as useful molecular tools for dissecting the relationship between the sequence of EGF ligand and its affect on receptor activation and signaling properties. Importantly, a handful of the EGF variants were rationally designed to have physicochemical profiles that are promising for their application as protein therapeutics. Finally, the toolkit of engineered cystine-rich proteins developed here can be used to glean future insight into the sequence-structure-function relationships of these two proteins and will inform future protein engineering efforts.
|Type of resource
|electronic; electronic resource; remote
|1 online resource.
|Lahti, Jennifer Lynn
|Stanford University, Department of Bioengineering.
|Cochran, Jennifer R
|Cochran, Jennifer R
|Statement of responsibility
|Jennifer Lynn Lahti.
|Submitted to the Department of Bioengineering.
|Thesis (Ph.D.)--Stanford University, 2010.
- © 2010 by Jennifer Lynn Lahti
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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