X-RAY STRUCTURE OF NUCLEOSIDE DIPHOSPHATE KINASE COMPLEXED WITH THYMIDINE DIPHOSPHATE AND MG2+ AT 2-ANGSTROM RESOLUTION

被引:74
作者
CHERFILS, J
MORERA, S
LASCU, I
VERON, M
JANIN, J
机构
[1] UNIV PARIS SUD,STRUCT BIOL LAB,CNRS,UMR 9920,F-91198 GIF SUR YVETTE,FRANCE
[2] UNIV BORDEAUX 2,CNRS,INST BIOCHIM CELLULAIRE,F-33077 BORDEAUX,FRANCE
[3] INST PASTEUR,UNITE BIOCHIM CELLULAIRE,CNRS,URA 1129,F-75724 PARIS 15,FRANCE
关键词
D O I
10.1021/bi00197a006
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We report the crystal structure of nucleoside diphosphate kinase (NDP kinase) from Dictyostelium discoideum with thymidine diphosphate (dTDP) and Mg2+ bound at the active site. The structure has been refined to an R-factor of 18.3% at 2-Angstrom resolution. The base stacks on the aromatic ring of Phe 64 near the protein surface and is wedged between the side chains of Phe 64 and Val 116. The sugar and the pyrophosphate are deeper inside the protein and make numerous H-bonds with protein side chains. There is no backbone interaction with the nucleotide. A Mg2+ ion bridges the alpha- and beta-phosphates and interacts with the protein via water molecules. NDP kinase shows little specificity toward ribonucleotides and deoxyribonucleotides. This property, required by the enzyme biological function, can now be analyzed by comparing the crystal structures of free, ADP-ligated, and dTDP-ligated enzymes. The most significant differences are located in residues 60-64, which adapt their conformation to allow Phe 64 to stack on both types of bases. Nonspecific binding is achieved by the absence of polar interaction between the base and protein atoms. The ribose of ADP and the deoxyribose of dTDP occupy similar positions, their hydroxyl groups interacting with Lys 16 and Asn 119. The H-bond between Lys 16 and the O-2' hydroxyl of ADP is replaced by a similar interaction with a water molecule in the dTDP complex. The beta-phosphate position is the same for ADP and dTDP, suggesting that the mechanism of phosphate transfer is the same for all substrates of NDP kinase. The NDP kinase binding site is compared with other nucleotide binding proteins with low specificity toward the base and sugar. We discuss DNA binding to NDP kinase and the design of nucleotide analogues which might be better substrates of NDP kinase than those currently used as antiviral agents.
引用
收藏
页码:9062 / 9069
页数:8
相关论文
共 43 条
  • [1] STRUCTURAL BASIS FOR THE 3'-5' EXONUCLEASE ACTIVITY OF ESCHERICHIA-COLI DNA-POLYMERASE-I - A 2 METAL-ION MECHANISM
    BEESE, LS
    STEITZ, TA
    [J]. EMBO JOURNAL, 1991, 10 (01) : 25 - 33
  • [2] A DROSOPHILA GENE THAT IS HOMOLOGOUS TO A MAMMALIAN GENE ASSOCIATED WITH TUMOR-METASTASIS CODES FOR A NUCLEOSIDE DIPHOSPHATE KINASE
    BIGGS, J
    HERSPERGER, E
    STEEG, PS
    LIOTTA, LA
    SHEARN, A
    [J]. CELL, 1990, 63 (05) : 933 - 940
  • [3] PHOSPHOTRANSFERASE AND SUBSTRATE BINDING MECHANISM OF THE CAMP-DEPENDENT PROTEIN-KINASE CATALYTIC SUBUNIT FROM PORCINE HEART AS DEDUCED FROM THE 2.0 ANGSTROM STRUCTURE OF THE COMPLEX WITH MN2+ ADENYLYL IMIDODIPHOSPHATE AND INHIBITOR PEPTIDE PKI(5-24)
    BOSSEMEYER, D
    ENGH, RA
    KINZEL, V
    PONSTINGL, H
    HUBER, R
    [J]. EMBO JOURNAL, 1993, 12 (03) : 849 - 859
  • [4] CRYSTALLOGRAPHIC R-FACTOR REFINEMENT BY MOLECULAR-DYNAMICS
    BRUNGER, AT
    KURIYAN, J
    KARPLUS, M
    [J]. SCIENCE, 1987, 235 (4787) : 458 - 460
  • [5] CRYSTAL-STRUCTURE OF THE AWD NUCLEOTIDE DIPHOSPHATE KINASE FROM DROSOPHILA
    CHIADMI, M
    MORERA, S
    LASCU, I
    DUMAS, C
    LEBRAS, G
    VERON, M
    JANIN, J
    [J]. STRUCTURE, 1993, 1 (04) : 283 - 293
  • [6] STAPHYLOCOCCAL NUCLEASE - PROPOSED MECHANISM OF ACTION BASED ON STRUCTURE OF ENZYME-THYMIDINE 3',5'-BISPHOSPHATE-CALCIUM ION COMPLEX AT 1.5-A RESOLUTION
    COTTON, FA
    HAZEN, EE
    LEGG, MJ
    [J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1979, 76 (06) : 2551 - 2555
  • [7] CROWTHER RA, 1972, MOL REPLACEMENT METH
  • [8] HIV INHIBITORS TARGETED AT THE REVERSE-TRANSCRIPTASE
    DECLERCQ, E
    [J]. AIDS RESEARCH AND HUMAN RETROVIRUSES, 1992, 8 (02) : 119 - 137
  • [9] X-RAY STRUCTURE OF NUCLEOSIDE DIPHOSPHATE KINASE
    DUMAS, C
    LASCU, I
    MORERA, S
    GLASER, P
    FOURME, R
    WALLET, V
    LACOMBE, ML
    VERON, M
    JANIN, J
    [J]. EMBO JOURNAL, 1992, 11 (09) : 3203 - 3208
  • [10] EALICK SE, 1990, J BIOL CHEM, V265, P1812