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Structural basis for SENP2 protease interactions with SUMO precursors and conjugated substrates

Nature Structural & Molecular Biology volume 13, pages 1060–1068 (2006)Cite this article

Abstract

SUMO processing and deconjugation are essential proteolytic activities for nuclear metabolism and cell-cycle progression in yeast and higher eukaryotes. To elucidate the mechanisms used during substrate lysine deconjugation, SUMO isoform processing and SUMO isoform interactions, X-ray structures were determined for a catalytically inert SENP2 protease domain in complex with conjugated RanGAP1–SUMO-1 or RanGAP1–SUMO-2, or in complex with SUMO-2 or SUMO-3 precursors. Common features within the active site include a 90° kink proximal to the scissile bond that forces C-terminal amino acid residues or the lysine side chain toward a protease surface that appears optimized for lysine deconjugation. Analysis of this surface reveals SENP2 residues, particularly Met497, that mediate, and in some instances reverse, in vitro substrate specificity. Mutational analysis and biochemistry provide a mechanism for SENP2 substrate preferences that explains why SENP2 catalyzes SUMO deconjugation more efficiently than processing.

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Figure 1: Structures of SENP2 deconjugation complexes with RanGAP1–SUMO-1 and RanGAP1–SUMO-2.
Figure 2: Comparative analysis of SENP2–SUMO-1 and SENP2–SUMO-2/3.
Figure 3: Structure of the SENP2–SUMO precursor complexes.
Figure 4: Structures of the productive and nonproductive SENP2–pre-SUMO-3 complexes.
Figure 5: Comparison of the hydrogen bond coordination of the ε and α peptide bonds in the deconjugation and processing complexes.
Figure 6: Comparative proteolysis assays for SUMO-1, SUMO-2 and SUMO-3 processing and deconjugation by SENP2 alanine-substituted mutants.
Figure 7: Comparative SENP2 mutagenesis analysis of processing reactions for pre-SUMO-1, pre-SUMO-2 and pre-SUMO-3 and deconjugation of RanGAP1–SUMO-1 and RanGAP1–SUMO-2/3 substrates.

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Acknowledgements

Use of the APS was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, under contract no. W-31-109-Eng-38. Use of the SGX Collaborative Access Team (SGX-CAT) beamline facilities at Sector 31 of the APS was provided by SGX Pharmaceuticals, Inc., who constructed and operates the facility. Use of the NE-CAT beamline at Sector 24 is based upon research conducted at the Northeastern Collaborative Access Team beamlines of the APS, which is supported by award RR-15301 from the National Center for Research Resources at the US National Institutes of Health (NIH). Beamline X29A at the National Synchrotron Light Source is supported by the Offices of Biological and Environmental Research and of Basic Energy Sciences of the US Department of Energy and the National Center for Research Resources of the NIH. D.R. and C.D.L. are supported in part by NIH grant GM65872. D.R. acknowledges support from the Charles H. Revson Foundation and C.D.L. acknowledges support from the Rita Allen Foundation.

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Authors and Affiliations

  1. Structural Biology Program, Sloan-Kettering Institute, New York, 10021, New York, USA

    David Reverter & Christopher D Lima

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  1. David Reverter
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D.R. generated the data. D.R. and C.D.L interpreted the data and wrote the manuscript.

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Correspondence to Christopher D Lima.

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Supplementary information

Supplementary Fig. 1

Sequence alignment for Senp/Ulp and SUMO family members. (PDF 67 kb)

Supplementary Fig. 2

Domain-swapped RanGAP1 structures. (PDF 630 kb)

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Reverter, D., Lima, C. Structural basis for SENP2 protease interactions with SUMO precursors and conjugated substrates. Nat Struct Mol Biol 13, 1060–1068 (2006). https://doi.org/10.1038/nsmb1168

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