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Prediction and assignment of function for a divergent N-succinyl amino acid racemase

Nature Chemical Biology volume 3, pages 486–491 (2007)Cite this article

Abstract

The protein databases contain many proteins with unknown function. A computational approach for predicting ligand specificity that requires only the sequence of the unknown protein would be valuable for directing experiment-based assignment of function. We focused on a family of unknown proteins in the mechanistically diverse enolase superfamily and used two approaches to assign function: (i) enzymatic assays using libraries of potential substrates, and (ii) in silico docking of the same libraries using a homology model based on the most similar (35% sequence identity) characterized protein. The results matched closely; an experimentally determined structure confirmed the predicted structure of the substrate-liganded complex. We assigned the N-succinyl arginine/lysine racemase function to the family, correcting the annotation (L-Ala-D/L-Glu epimerase) based on the function of the most similar characterized homolog. These studies establish that ligand docking to a homology model can facilitate functional assignment of unknown proteins by restricting the identities of the possible substrates that must be experimentally tested.

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Figure 1: Structure, functions and sequence relationships in the MLE subgroup.
Figure 2: Active sites of the template AEE and the modeled BC0371.
Figure 3: Active site of BC0371 complexed with N-succinyl-L-arginine as predicted by homology modeling and experimentally determined by X-ray crystallography.

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Acknowledgements

This work was supported by the US National Institutes of Health (1 P01 GM-71790).

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Authors and Affiliations

  1. Departments of Biochemistry and Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, 61801, Illinois, USA

    Ling Song, Heidi J Imker & John A Gerlt

  2. Department of Pharmaceutical Chemistry, School of Pharmacy, University of California, 600 16th Street, San Francisco, 94158, California, USA

    Chakrapani Kalyanaraman, Patricia C Babbitt & Matthew P Jacobson

  3. Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, 10461, New York, USA

    Alexander A Fedorov, Elena V Fedorov & Steven C Almo

  4. Department of Biopharmaceutical Sciences, School of Pharmacy, University of California, 1700 4th Street, San Francisco, 94158, California, USA

    Margaret E Glasner, Shoshana Brown & Patricia C Babbitt

  5. California Institute for Quantitative Biomedical Research, University of California, 1700 4th Street, San Francisco, 94158, California, USA

    Patricia C Babbitt

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  1. Ling Song
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Contributions

L.S. isolated BC0371 and performed library screening and kinetic assays; C.K. performed molecular modeling and in silico library docking; A.A.F. and E.V.F. crystallized BC0371 and solved its structure; M.E.G. and S.B. performed sequence alignments and generated the Cytoscape-based representation of sequence relationships; H.J.I. cloned the gene encoding BC0371 and spectroscopically characterized dipeptide substrates; P.C.B., S.C.A., M.P.J. and J.A.G. designed the experiments and wrote the manuscript.

Corresponding authors

Correspondence to Matthew P Jacobson or John A Gerlt.

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M.P.J. is a member of the Scientific Advisory Board of Schrodinger Inc.

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Supplementary Figures 1–3, Supplementary Table 1 and Supplementary Methods (PDF 1201 kb)

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Song, L., Kalyanaraman, C., Fedorov, A. et al. Prediction and assignment of function for a divergent N-succinyl amino acid racemase. Nat Chem Biol 3, 486–491 (2007). https://doi.org/10.1038/nchembio.2007.11

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