PDBsum entry 1lwt

Go to PDB code: 
protein dna_rna links
Hydrolase/DNA PDB id
Protein chain
448 a.a. *
Waters ×15
* Residue conservation analysis
PDB id:
Name: Hydrolase/DNA
Title: Crystal structure of the intein homing endonuclease pi-scei bound to its substrate DNA (ca2+ free)
Structure: Pi-scei DNA substrate top strand. Chain: b. Engineered: yes. Pi-scei DNA substrate bottom strand. Chain: c. Engineered: yes. Endonuclease pi-scei. Chain: a. Synonym: pi-scei, vma-derived endonuclease, vde, sce vma
Source: Synthetic: yes. Saccharomyces cerevisiae. Baker's yeast. Organism_taxid: 4932. Gene: vma1. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Trimer (from PQS)
3.20Å     R-factor:   0.242     R-free:   0.287
Authors: C.M.Moure,F.S.Gimble,F.A.Quiocho
Key ref:
C.M.Moure et al. (2002). Crystal structure of the intein homing endonuclease PI-SceI bound to its recognition sequence. Nat Struct Biol, 9, 764-770. PubMed id: 12219083 DOI: 10.1038/nsb840
03-Jun-02     Release date:   27-Sep-02    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P17255  (VATA_YEAST) -  V-type proton ATPase catalytic subunit A
1071 a.a.
448 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.  - H(+)-transporting two-sector ATPase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: ATP + H2O + H+(In) = ADP + phosphate + H+(Out)
+ H(2)O
+ H(+)(In)
+ phosphate
+ H(+)(Out)
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     protein splicing   2 terms 
  Biochemical function     DNA binding     2 terms  


DOI no: 10.1038/nsb840 Nat Struct Biol 9:764-770 (2002)
PubMed id: 12219083  
Crystal structure of the intein homing endonuclease PI-SceI bound to its recognition sequence.
C.M.Moure, F.S.Gimble, F.A.Quiocho.
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 Ca(2+). The DNA shows an asymmetric bending pattern, with a major 50 degree bend in the endonuclease domain and a minor 22 degree 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.
  Selected figure(s)  
Figure 4.
Figure 4. PI-SceI-DNA contacts in the orthorhombic structure. a, Schematic diagram. Only polar contacts (<3.5 ) are shown. Upstream (negative numbers) and downstream bases (positive numbers) are blue and green, respectively. Interactions involving the splicing and endonuclease domains are dark blue and red, respectively. The two scissile phosphates are indicated by a filled circle. Minor and major groove interactions are indicated with dashed and solid lines, respectively. b, Stereo view of the protein-DNA interface showing the antiparallel strands 21- 22 and Lys 340 (a 20 residue) interacting with the DNA. This region induces a 28 roll in the DNA at the +4/+3 step. c, Stereo view of the protein -DNA interface in the DRR region. The DRR region stabilizes a 22 roll in the DNA at the +18/+19 step.
Figure 6.
Figure 6. Stereo view of the superimposition of the PI-SceI (hexagonal structure) and I-CreI catalytic centers with bound DNA. The superimposition (r.m.s. deviation of 3.7 ) was achieved by matching similar secondary structure elements in the endonuclease domain of PI-SceI (cyan) and the I-CreI (yellow and brown) homodimer22 (PDB entry 1G9Y). The overlapped active sites residues are labeled in black (PI-SceI) and red (I-CreI). The positions of the scissile phosphates in I-CreI and PI-SceI are marked with orange spheres on DNA strands colored gray and green, respectively. The bound Ca^2+ ions are indicated by smaller yellow (I-CreI) or dark blue (PI-SceI) spheres. Lys 301 and its pseudo symmetric pair Lys 403 superimpose with Lys 98 and Lys 98' in I-CreI, which are engaged in the formation of the hydration network (not shown) at the catalytic center. Asp 229 and Thr 341 superimpose with Gln 47 and Gln 47', respectively, of I-CreI which also bind water molecules. The importance of the PI-SceI residues in catalysis is supported by mutagenesis studies7, 24, 36.
  The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Biol (2002, 9, 764-770) copyright 2002.  
  Figures were selected by the author.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21220111 B.L.Stoddard (2011).
Homing endonucleases: from microbial genetic invaders to reagents for targeted DNA modification.
  Structure, 19, 7.  
21047873 S.Arnould, C.Delenda, S.Grizot, C.Desseaux, F.Pâques, G.H.Silva, and J.Smith (2011).
The I-CreI meganuclease and its engineered derivatives: applications from cell modification to gene therapy.
  Protein Eng Des Sel, 24, 27-31.  
19915993 M.J.Marcaida, I.G.Muñoz, F.J.Blanco, J.Prieto, and G.Montoya (2010).
Homing endonucleases: from basics to therapeutic applications.
  Cell Mol Life Sci, 67, 727-748.  
19937653 P.Singh, P.Tripathi, and K.Muniyappa (2010).
Mutational analysis of active-site residues in the Mycobacterium leprae RecA intein, a LAGLIDADG homing endonuclease: Asp(122) and Asp(193) are crucial to the double-stranded DNA cleavage activity whereas Asp(218) is not.
  Protein Sci, 19, 111-123.  
19913037 S.E.Lindner, E.K.De Silva, J.L.Keck, and M.Llinás (2010).
Structural determinants of DNA binding by a P. falciparum ApiAP2 transcriptional regulator.
  J Mol Biol, 395, 558-567.
PDB code: 3igm
19153140 H.Li, S.Pellenz, U.Ulge, B.L.Stoddard, and R.J.Monnat (2009).
Generation of single-chain LAGLIDADG homing endonucleases from native homodimeric precursor proteins.
  Nucleic Acids Res, 37, 1650-1662.
PDB code: 3fd2
19605345 P.Singh, P.Tripathi, G.H.Silva, A.Pingoud, and K.Muniyappa (2009).
Characterization of Mycobacterium leprae RecA intein, a LAGLIDADG homing endonuclease, reveals a unique mode of DNA binding, helical distortion, and cleavage compared with a canonical LAGLIDADG homing endonuclease.
  J Biol Chem, 284, 25912-25928.  
18424798 C.M.Moure, F.S.Gimble, and F.A.Quiocho (2008).
Crystal structures of I-SceI complexed to nicked DNA substrates: snapshots of intermediates along the DNA cleavage reaction pathway.
  Nucleic Acids Res, 36, 3287-3296.
PDB codes: 3c0w 3c0x
18276641 E.Fajardo-Sanchez, F.Stricher, F.Pâques, M.Isalan, and L.Serrano (2008).
Computer design of obligate heterodimer meganucleases allows efficient cutting of custom DNA sequences.
  Nucleic Acids Res, 36, 2163-2173.  
18974222 M.J.Marcaida, J.Prieto, P.Redondo, A.D.Nadra, A.Alibés, L.Serrano, S.Grizot, P.Duchateau, F.Pâques, F.J.Blanco, and G.Montoya (2008).
Crystal structure of I-DmoI in complex with its target DNA provides new insights into meganuclease engineering.
  Proc Natl Acad Sci U S A, 105, 16888-16893.
PDB codes: 2vs7 2vs8
18644379 Y.Niu, K.Tenney, H.Li, and F.S.Gimble (2008).
Engineering variants of the I-SceI homing endonuclease with strand-specific and site-specific DNA-nicking activity.
  J Mol Biol, 382, 188-202.  
  17389927 A.G.Senejani, and J.P.Gogarten (2007).
Structural stability and endonuclease activity of a PI-SceI GFP-fusion protein.
  Int J Biol Sci, 3, 205-211.  
17586768 M.A.Johnson, M.W.Southworth, T.Herrmann, L.Brace, F.B.Perler, and K.Wüthrich (2007).
NMR structure of a KlbA intein precursor from Methanococcus jannaschii.
  Protein Sci, 16, 1316-1328.
PDB codes: 2jmz 2jnq
17720189 M.Scalley-Kim, A.McConnell-Smith, and B.L.Stoddard (2007).
Coevolution of a homing endonuclease and its host target sequence.
  J Mol Biol, 372, 1305-1319.
PDB code: 2qoj
17426121 P.Volná, J.Jarjour, S.Baxter, S.R.Roffler, R.J.Monnat, B.L.Stoddard, and A.M.Scharenberg (2007).
Flow cytometric analysis of DNA binding and cleavage by cell surface-displayed homing endonucleases.
  Nucleic Acids Res, 35, 2748-2758.  
16507575 A.Bakhrat, K.Baranes, O.Krichevsky, I.Rom, G.Schlenstedt, S.Pietrokovski, and D.Raveh (2006).
Nuclear import of ho endonuclease utilizes two nuclear localization signals and four importins of the ribosomal import system.
  J Biol Chem, 281, 12218-12226.  
16493661 H.Matsumura, H.Takahashi, T.Inoue, T.Yamamoto, H.Hashimoto, M.Nishioka, S.Fujiwara, M.Takagi, T.Imanaka, and Y.Kai (2006).
Crystal structure of intein homing endonuclease II encoded in DNA polymerase gene from hyperthermophilic archaeon Thermococcus kodakaraensis strain KOD1.
  Proteins, 63, 711-715.
PDB codes: 2cw7 2cw8
17101053 J.P.Gogarten, and E.Hilario (2006).
Inteins, introns, and homing endonucleases: recent revelations about the life cycle of parasitic genetic elements.
  BMC Evol Biol, 6, 94.  
16698548 P.C.Spiegel, B.Chevalier, D.Sussman, M.Turmel, C.Lemieux, and B.L.Stoddard (2006).
The structure of I-CeuI homing endonuclease: Evolving asymmetric DNA recognition from a symmetric protein scaffold.
  Structure, 14, 869-880.
PDB code: 2ex5
16049020 N.Nomura, Y.Morinaga, N.Shirai, and Y.Sako (2005).
I-ApeKI [corrected]: a novel intron-encoded LAGLIDADG homing endonuclease from the archaeon, Aeropyrum pernix K1.
  Nucleic Acids Res, 33, e116.  
16040597 S.Balaji, M.M.Babu, L.M.Iyer, and L.Aravind (2005).
Discovery of the principal specific transcription factors of Apicomplexa and their implication for the evolution of the AP2-integrase DNA binding domains.
  Nucleic Acids Res, 33, 3994-4006.  
15862101 T.C.Evans, M.Q.Xu, and S.Pradhan (2005).
Protein splicing elements and plants: from transgene containment to protein purification.
  Annu Rev Plant Biol, 56, 375-392.  
15020462 A.Bakhrat, M.S.Jurica, B.L.Stoddard, and D.Raveh (2004).
Homology modeling and mutational analysis of Ho endonuclease of yeast.
  Genetics, 166, 721-728.  
15531154 A.Burt, and V.Koufopanou (2004).
Homing endonuclease genes: the rise and fall and rise again of a selfish element.
  Curr Opin Genet Dev, 14, 609-615.  
14634013 A.J.Noël, W.Wende, and A.Pingoud (2004).
DNA recognition by the homing endonuclease PI-SceI involves a divalent metal ion cofactor-induced conformational change.
  J Biol Chem, 279, 6794-6804.  
14745027 G.Butler, C.Kenny, A.Fagan, C.Kurischko, C.Gaillardin, and K.H.Wolfe (2004).
Evolution of the MAT locus and its Ho endonuclease in yeast species.
  Proc Natl Acad Sci U S A, 101, 1632-1637.  
15190132 G.H.Silva, and M.Belfort (2004).
Analysis of the LAGLIDADG interface of the monomeric homing endonuclease I-DmoI.
  Nucleic Acids Res, 32, 3156-3168.  
14962381 K.C.Hsia, K.F.Chak, P.H.Liang, Y.S.Cheng, W.Y.Ku, and H.S.Yuan (2004).
DNA binding and degradation by the HNH protein ColE7.
  Structure, 12, 205-214.
PDB code: 1pt3
15280510 K.L.Posey, V.Koufopanou, A.Burt, and F.S.Gimble (2004).
Evolution of divergent DNA recognition specificities in VDE homing endonucleases from two yeast species.
  Nucleic Acids Res, 32, 3947-3956.  
14764082 R.David, M.P.Richter, and A.G.Beck-Sickinger (2004).
Expressed protein ligation. Method and applications.
  Eur J Biochem, 271, 663-677.  
14760742 S.Steuer, V.Pingoud, A.Pingoud, and W.Wende (2004).
Chimeras of the homing endonuclease PI-SceI and the homologous Candida tropicalis intein: a study to explore the possibility of exchanging DNA-binding modules to obtain highly specific endonucleases with altered specificity.
  Chembiochem, 5, 206-213.  
15502310 T.Imagawa, H.Nakayama, N.Katunuma, H.Sakuraba, T.Ohshima, T.Itoh, Y.Sako, N.Nomura, and H.Tsuge (2004).
Crystallization and preliminary X-ray diffraction analysis of homing endonuclease I-Tsp061I.
  Acta Crystallogr D Biol Crystallogr, 60, 2006-2008.  
12750473 G.Sasnauskas, S.E.Halford, and V.Siksnys (2003).
How the BfiI restriction enzyme uses one active site to cut two DNA strands.
  Proc Natl Acad Sci U S A, 100, 6410-6415.  
12771221 J.C.Epinat, S.Arnould, P.Chames, P.Rochaix, D.Desfontaines, C.Puzin, A.Patin, A.Zanghellini, F.Pâques, and E.Lacroix (2003).
A novel engineered meganuclease induces homologous recombination in yeast and mammalian cells.
  Nucleic Acids Res, 31, 2952-2962.  
14633971 J.M.Bolduc, P.C.Spiegel, P.Chatterjee, K.L.Brady, M.E.Downing, M.G.Caprara, R.B.Waring, and B.L.Stoddard (2003).
Structural and biochemical analyses of DNA and RNA binding by a bifunctional homing endonuclease and group I intron splicing factor.
  Genes Dev, 17, 2875-2888.
PDB code: 1p8k
12853636 N.Guhan, and K.Muniyappa (2003).
Mycobacterium tuberculosis RecA intein, a LAGLIDADG homing endonuclease, displays Mn(2+) and DNA-dependent ATPase activity.
  Nucleic Acids Res, 31, 4184-4191.  
12654010 W.J.Geese, Y.K.Kwon, X.Wen, and R.B.Waring (2003).
In vitro analysis of the relationship between endonuclease and maturase activities in the bi-functional group I intron-encoded protein, I-AniI.
  Eur J Biochem, 270, 1543-1554.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.