Uma sequência de código mínima para mudar a estrutura e a função da proteína

A minimal sequence code for switching protein structure and function

Patrick A. Alexander, Yanan He, Yihong Chen, John Orban and Philip N. Bryan1

+ Author Affiliations

Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville, MD 20850
Edited by Adriaan Bax, National Institutes of Health, Bethesda, MD, and approved October 12, 2009 (received for review June 9, 2009)

Abstract

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, 4β+α fold can be transformed into an albumin-binding, 3-α 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 4β+α to 3-α structure can occur via a single amino acid substitution. On one side of the switch point, the 4β+α fold is >90% populated (pH 7.2, 20 °C). A single mutation switches the conformation to the 3-α fold, which is >90% populated (pH 7.2, 20 °C). We further show that a bifunctional protein exists at the switch point with affinity for both IgG and albumin.

evolution NMR protein design protein folding

Footnotes

1To whom correspondence should be addressed. E-mail: bryan@umbi.umd.edu

Author contributions: P.A.A., Y.H., J.O., and P.N.B. designed research; P.A.A., Y.H., Y.C., J.O., and P.N.B. performed research; P.A.A., Y.H., J.O., and P.N.B. analyzed data; and P.N.B. wrote the paper.

The authors declare no conflict of interest.

This article is a PNAS Direct Submission.

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