A model for the mechanism of human topoisomerase I.

The three-dimensional structure of a 70-kilodalton amino terminally truncated form of human topoisomerase I in complex with a 22-base pair duplex oligonucleotide, determined to a resolution of 2.8 angstroms, reveals all of the structural elements of the enzyme that contact DNA. The linker region that connects the central core of the enzyme to the carboxyl-terminal domain assumes a coiled-coil configuration and protrudes away from the remainder of the enzyme. The positively charged DNA-proximal surface of the linker makes only a few contacts with the DNA downstream of the cleavage site. In combination with the crystal structures of the reconstituted human topoisomerase I before and after DNA cleavage, this information suggests which amino acid residues are involved in catalyzing phosphodiester bond breakage and religation. The structures also lead to the proposal that the topoisomerization step occurs by a mechanism termed "controlled rotation."

[1]  Alfonso Mondragón,et al.  Three-dimensional structure of the 67K N-terminal fragment of E. coli DNA topoisomerase I , 1994, Nature.

[2]  E. Fauman,et al.  Crystal structure of Yersinia protein tyrosine phosphatase at 2.5 Å and the complex with tungstate , 1994, Nature.

[3]  P. Brown,et al.  A sign inversion mechanism for enzymatic supercoiling of DNA. , 1979, Science.

[4]  S. Darst,et al.  Crystal structure of the GreA transcript cleavage factor from Escherichia coli , 1995, Nature.

[5]  P. Wright,et al.  1H NMR studies of plastocyanin from Scenedesmus obliquus: complete sequence-specific assignment, secondary structure analysis, and global fold. , 1988, Biochemistry.

[6]  S. Shuman,et al.  Mutational analysis of vaccinia DNA topoisomerase defines amino acid residues essential for covalent catalysis. , 1994, The Journal of biological chemistry.

[7]  D. Barford,et al.  Crystal structure of human protein tyrosine phosphatase 1B. , 1994, Science.

[8]  F. Guo,et al.  Structure of Cre recombinase complexed with DNA in a site-specific recombination synapse , 1997, Nature.

[9]  D. Esposito,et al.  The integrase family of tyrosine recombinases: evolution of a conserved active site domain. , 1997, Nucleic acids research.

[10]  Axel T. Brunger,et al.  X-PLOR Version 3.1: A System for X-ray Crystallography and NMR , 1992 .

[11]  A. Andersen,et al.  Topoisomerase I has a strong binding preference for a conserved hexadecameric sequence in the promoter region of the rRNA gene from Tetrahymena pyriformis. , 1985, Nucleic acids research.

[12]  P. Kraulis.  A program to produce both detailed and schematic plots of protein structures , 1991 .

[13]  M. Bjornsti,et al.  Alterations in the Catalytic Activity of Yeast DNA Topoisomerase I Result in Cell Cycle Arrest and Cell Death* , 1997, The Journal of Biological Chemistry.

[14]  G. Fink,et al.  A novel mutation in DNA topoisomerase I of yeast causes DNA damage and RAD9-dependent cell cycle arrest. , 1993, Genetics.

[15]  Chris Sander,et al.  Dali/FSSP classification of three-dimensional protein folds , 1997, Nucleic Acids Res..

[16]  T. Ellenberger,et al.  Flexibility in DNA Recombination: Structure of the Lambda Integrase Catalytic Core , 1997, Science.

[17]  A. Brunger.  Crystallographic refinement by simulated annealing , 1988 .

[18]  J. Svejstrup,et al.  Purification and characterization of human topoisomerase I mutants. , 1996, European journal of biochemistry.

[19]  G. Ireton,et al.  The Domain Organization of Human Topoisomerase I (*) , 1996, The Journal of Biological Chemistry.

[20]  J. Champoux,et al.  Overexpression of human topoisomerase I in baby hamster kidney cells: hypersensitivity of clonal isolates to camptothecin. , 1992, Cancer research.

[21]  F. Dyda,et al.  Molecular Organization in Site-Specific Recombination: The Catalytic Domain of Bacteriophage HP1 Integrase at 2.7 Å Resolution , 1997, Cell.

[22]  D. Tsernoglou,et al.  Restored heptad pattern continuity does not alter the folding of a four-α-helix bundle , 1994, Nature Structural Biology.

[23]  J. Champoux,et al.  The role of single-strand breaks in the catenation reaction catalyzed by the rat type I topoisomerase. , 1986, The Journal of biological chemistry.

[24]  T. Harris,et al.  Vaccinia DNA topoisomerase I: evidence supporting a free rotation mechanism for DNA supercoil relaxation. , 1997, Biochemistry.

[25]  Nicholas R. Cozzarelli,et al.  DNA topology and its biological effects , 1990 .