A minimal sequence code for switching protein structure and function.

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TitleA minimal sequence code for switching protein structure and function.
Publication TypeJournal Article
Year of Publication2009
AuthorsAlexander, PA, He, Y, Chen, Y, Orban, J, Bryan, PN
JournalProc Natl Acad Sci U S A
Date Published2009 Dec 15
KeywordsHumans, Immunoglobulin G, Magnetic Resonance Spectroscopy, Mutagenesis, Site-Directed, Mutation, Protein Conformation, Protein Engineering, Protein Folding, Protein Stability, Recombinant Fusion Proteins, Serum Albumin, Subtilisin

We present here a structural and mechanistic description of how a protein changes its fold and function, mutation by mutation. Our approach was to create 2 proteins that (i) are stably folded into 2 different folds, (ii) have 2 different functions, and (iii) are very similar in sequence. In this simplified sequence space we explore the mutational path from one fold to another. We show that an IgG-binding, 4beta+alpha fold can be transformed into an albumin-binding, 3-alpha fold via a mutational pathway in which neither function nor native structure is completely lost. The stabilities of all mutants along the pathway are evaluated, key high-resolution structures are determined by NMR, and an explanation of the switching mechanism is provided. We show that the conformational switch from 4beta+alpha to 3-alpha structure can occur via a single amino acid substitution. On one side of the switch point, the 4beta+alpha fold is >90% populated (pH 7.2, 20 degrees C). A single mutation switches the conformation to the 3-alpha fold, which is >90% populated (pH 7.2, 20 degrees C). We further show that a bifunctional protein exists at the switch point with affinity for both IgG and albumin.

Alternate JournalProc. Natl. Acad. Sci. U.S.A.
PubMed ID19923431
PubMed Central IDPMC2779201
Grant ListGM62154 / GM / NIGMS NIH HHS / United States