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      RosettaRemodel: A Generalized Framework for Flexible Backbone Protein Design

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          Abstract

          We describe RosettaRemodel, a generalized framework for flexible protein design that provides a versatile and convenient interface to the Rosetta modeling suite. RosettaRemodel employs a unified interface, called a blueprint, which allows detailed control over many aspects of flexible backbone protein design calculations. RosettaRemodel allows the construction and elaboration of customized protocols for a wide range of design problems ranging from loop insertion and deletion, disulfide engineering, domain assembly, loop remodeling, motif grafting, symmetrical units, to de novo structure modeling.

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          Most cited references 51

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          Protein structure prediction using Rosetta.

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            Kemp elimination catalysts by computational enzyme design.

            The design of new enzymes for reactions not catalysed by naturally occurring biocatalysts is a challenge for protein engineering and is a critical test of our understanding of enzyme catalysis. Here we describe the computational design of eight enzymes that use two different catalytic motifs to catalyse the Kemp elimination-a model reaction for proton transfer from carbon-with measured rate enhancements of up to 10(5) and multiple turnovers. Mutational analysis confirms that catalysis depends on the computationally designed active sites, and a high-resolution crystal structure suggests that the designs have close to atomic accuracy. Application of in vitro evolution to enhance the computational designs produced a >200-fold increase in k(cat)/K(m) (k(cat)/K(m) of 2,600 M(-1)s(-1) and k(cat)/k(uncat) of >10(6)). These results demonstrate the power of combining computational protein design with directed evolution for creating new enzymes, and we anticipate the creation of a wide range of useful new catalysts in the future.
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              De novo computational design of retro-aldol enzymes.

              The creation of enzymes capable of catalyzing any desired chemical reaction is a grand challenge for computational protein design. Using new algorithms that rely on hashing techniques to construct active sites for multistep reactions, we designed retro-aldolases that use four different catalytic motifs to catalyze the breaking of a carbon-carbon bond in a nonnatural substrate. Of the 72 designs that were experimentally characterized, 32, spanning a range of protein folds, had detectable retro-aldolase activity. Designs that used an explicit water molecule to mediate proton shuffling were significantly more successful, with rate accelerations of up to four orders of magnitude and multiple turnovers, than those involving charged side-chain networks. The atomic accuracy of the design process was confirmed by the x-ray crystal structure of active designs embedded in two protein scaffolds, both of which were nearly superimposable on the design model.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2011
                31 August 2011
                : 6
                : 8
                Affiliations
                [1 ]Department of Biochemistry, University of Washington, Seattle, Washington, United States of America
                [2 ]Interdisciplinary Program in Biomolecular Structure and Design, University of Washington, Seattle, Washington, United States of America
                [3 ]Department of Biochemistry and Structural Biology, Lund University, Lund, Sweden
                [4 ]Program in Molecular Structure and Function, Hospital for Sick Children, Toronto, Ontario, Canada
                [5 ]Howard Hughes Medical Institute, University of Washington, Seattle, Washington, United States of America
                University of South Florida College of Medicine, United States of America
                Author notes

                Conceived and designed the experiments: P-SH YAB WRS DB. Contributed reagents/materials/analysis tools: FR IA RV. Wrote the paper: P-SH YAB WRS DB.

                [¤a]

                Current address: Arzeda Corporation, Seattle, Washington, United States of America

                [¤b]

                Current address: IAVI Neutralizing Antibody Center and Department of Immunology and Microbial Sciences, The Scripps Research Institute, La Jolla, California, United States of America

                Article
                PONE-D-11-09414
                10.1371/journal.pone.0024109
                3166072
                21909381
                a395b748-6538-4057-9c10-0197419a523d
                Huang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                Page count
                Pages: 8
                Categories
                Research Article
                Biology
                Biochemistry
                Proteins
                Protein Structure
                Computational Biology
                Macromolecular Structure Analysis
                Protein Structure
                Proteomics
                Protein Engineering

                Uncategorized

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