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Crystal structure of the intein homing endonuclease PI-SceI bound to its recognition sequence

Nature Structural Biology volume 9, pages 764–770 (2002)Cite this article

Abstract

The first X-ray structures of an intein–DNA complex, that of the two-domain homing endonuclease PI-SceI bound to its 36-base pair DNA substrate, have been determined in the presence and absence of Ca2+. The DNA shows an asymmetric bending pattern, with a major 50° bend in the endonuclease domain and a minor 22° bend in the splicing domain region. Distortions of the DNA bound to the endonuclease domain cause the insertion of the two cleavage sites in the catalytic center. DNA binding induces changes in the protein conformation. The two overlapping non-identical active sites in the endonucleolytic center contain two Ca+2 ions that coordinate to the catalytic Asp residues. Structure analysis indicates that the top strand may be cleaved first.

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Figure 1: DNA substrate of PI-SceI.
Figure 2: Overview of the PI-SceI-DNA complex.
Figure 3: Structural parameters of the DNA bound in the orthorhombic structure.
Figure 4: PI-SceI-DNA contacts in the orthorhombic structure.
Figure 5: Superimposition of the DNA-bound structures of the PI-SceI endonuclease domain (cyan) and the symmetry-related I-CreI dimer22 (PDB entry 1G9Y) (red-orange and violet).
Figure 6: Stereo view of the superimposition of the PI-SceI (hexagonal structure) and I-CreI catalytic centers with bound DNA.

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Acknowledgements

This work was supported by the Welch Foundation grants to F.S.G. and to F.A.Q. and the Howard Hughes Medical Institute, with which C.M.M. is a Research Associate and F.A.Q. an Investigator. We thank N. Vyas for useful discussions and B. Bowman and the BIOCARS staff at the APS in Chicago, especially V. Srajer and P. Reinhard for their assistance in data collection.

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Authors and Affiliations

  1. Howard Hughes Medical Institute, Baylor College of Medicine, Houston, 77030, Texas, USA

    Carmen M. Moure & Florante A. Quiocho

  2. Institute of Biosciences and Technology and Department of Medical Biochemistry and Genetics, Center for Genome Research, Texas A&M University System Health Science Center, 2121 West Holcombe Boulevard, Houston, 77030, Texas, USA

    Frederick S. Gimble

  3. Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, 77030, Texas, USA

    Florante A. Quiocho

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Correspondence to Florante A. Quiocho.

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Moure, C., Gimble, F. & Quiocho, F. Crystal structure of the intein homing endonuclease PI-SceI bound to its recognition sequence. Nat Struct Mol Biol 9, 764–770 (2002). https://doi.org/10.1038/nsb840

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