From synthetic coiled coils to functional proteins: Automated design of a receptor for the calmodulin-binding domain of calcineurin

Giovanna Ghirlanda; James D. Lear; Angela Lombardi; William F. Degrado

doi:10.1006/jmbi.1998.1912

From synthetic coiled coils to functional proteins: Automated design of a receptor for the calmodulin-binding domain of calcineurin

Giovanna Ghirlanda, James D. Lear, Angela Lombardi, William F. Degrado

Arizona State University

Research output: Contribution to journal › Article › peer-review

37 Citations (Scopus)

Abstract

A series of synthetic receptors capable of binding to the calmodulin-binding domain of calcineurin (CN_393-414) was designed, synthesized and characterized. The design was accomplished by docking CN_393-414 against a two-helix receptor, using an idealized three-stranded coiled coil as a starting geometry. The sequence of the receptor was chosen using a side-chain re-packing program, which employed a genetic algorithm to select potential binders from a total of 7.5 x 10⁶ possible sequences. A total of 25 receptors were prepared, representing 13 sequences predicted by the algorithm as well as 12 related sequences that were not predicted. The receptors were characterized by CD spectroscopy, analytical ultracentrifugation, and binding assays. The receptors predicted by the algorithm bound CN_393-414 with apparent dissociation constants ranging from 0.2 μM to > 50 μM. Many of the receptors that were not predicted by the algorithm also bound to CN_393-414. Methods to circumvent this problem and to improve the automated design of functional proteins are discussed.

Original language	English
Pages (from-to)	379-391
Number of pages	13
Journal	Journal of Molecular Biology
Volume	281
Issue number	2
DOIs	https://doi.org/10.1006/jmbi.1998.1912
Publication status	Published - Aug 14 1998

Keywords

Calmodulin mimetic
De novo design
Molecular recognition
Peptide binding
Three-helix bundle

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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title = "From synthetic coiled coils to functional proteins: Automated design of a receptor for the calmodulin-binding domain of calcineurin",

abstract = "A series of synthetic receptors capable of binding to the calmodulin-binding domain of calcineurin (CN393-414) was designed, synthesized and characterized. The design was accomplished by docking CN393-414 against a two-helix receptor, using an idealized three-stranded coiled coil as a starting geometry. The sequence of the receptor was chosen using a side-chain re-packing program, which employed a genetic algorithm to select potential binders from a total of 7.5 x 106 possible sequences. A total of 25 receptors were prepared, representing 13 sequences predicted by the algorithm as well as 12 related sequences that were not predicted. The receptors were characterized by CD spectroscopy, analytical ultracentrifugation, and binding assays. The receptors predicted by the algorithm bound CN393-414 with apparent dissociation constants ranging from 0.2 μM to > 50 μM. Many of the receptors that were not predicted by the algorithm also bound to CN393-414. Methods to circumvent this problem and to improve the automated design of functional proteins are discussed.",

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author = "Giovanna Ghirlanda and Lear, {James D.} and Angela Lombardi and Degrado, {William F.}",

note = "Funding Information: The authors thank John Desjarlais and Tracy Handel for the use of the NBSEARCH and ROC1 programs, and S. Betz, G. Dieckmann, B. Hill, H. Lu, J. Schneider and C. Summa for many useful discussions. This work was supported by NIH grant GM54616 and in part by the MRSEC Program of the National Science Foundation under award number DMR96-32598, and NIH grant GM54616. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.",

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From synthetic coiled coils to functional proteins: Automated design of a receptor for the calmodulin-binding domain of calcineurin

Abstract

Keywords

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