1DFA
CRYSTAL STRUCTURE OF PI-SCEI IN C2 SPACE GROUP
Summary for 1DFA
| Entry DOI | 10.2210/pdb1dfa/pdb |
| Related | 1VDE |
| Descriptor | PI-SCEI ENDONUCLEASE (2 entities in total) |
| Functional Keywords | intein, homing endonuclease, hydrolase |
| Biological source | Saccharomyces cerevisiae (baker's yeast) |
| Cellular location | Endomembrane system: P17255 |
| Total number of polymer chains | 1 |
| Total formula weight | 51003.89 |
| Authors | Hu, D.,Crist, M.,Duan, X.,Quiocho, F.A.,Gimble, F.S. (deposition date: 1999-11-18, release date: 1999-12-08, Last modification date: 2024-02-07) |
| Primary citation | Hu, D.,Crist, M.,Duan, X.,Quiocho, F.A.,Gimble, F.S. Probing the structure of the PI-SceI-DNA complex by affinity cleavage and affinity photocross-linking. J.Biol.Chem., 275:2705-2712, 2000 Cited by PubMed Abstract: The PI-SceI protein is an intein-encoded homing endonuclease that initiates the mobility of its gene by making a double strand break at a single site in the yeast genome. The PI-SceI protein splicing and endonucleolytic active sites are separately located in each of two domains in the PI-SceI structure. To determine the spatial relationship between bases in the PI-SceI recognition sequence and selected PI-SceI amino acids, the PI-SceI-DNA complex was probed by photocross-linking and affinity cleavage methods. Unique solvent-accessible cysteine residues were introduced into the two PI-SceI domains at positions 91, 97, 170, 230, 376, and 378, and the mutant proteins were modified with either 4-azidophenacyl bromide or iron (S)-1-(p-bromoacetamidobenzyl)-ethylenediaminetetraacetate (FeBABE). The phenyl azide-coupled proteins cross-linked to the PI-SceI target sequence, and the FeBABE-modified proteins cleaved the DNA proximal to the derivatized amino acid. The results suggest that an extended beta-hairpin loop in the endonuclease domain that contains residues 376 and 378 contacts the major groove near the PI-SceI cleavage site. Conversely, residues 91, 97, and 170 in the protein splicing domain are in close proximity to a distant region of the substrate. To interpret our results, we used a new PI-SceI structure that is ordered in regions of the protein that bind DNA. The data strongly support a model of the PI-SceI-DNA complex derived from this structure. PubMed: 10644733DOI: 10.1074/jbc.275.4.2705 PDB entries with the same primary citation |
| Experimental method | X-RAY DIFFRACTION (2 Å) |
Structure validation
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