Solution Structure of Neocarzinostatin Determined by Homonuclear Two-Dimensional Nuclear Magnetic Resonance

  • Hiroyuki Takashima
  • Yuji Kobayashi
Chapter

Summary

The secondary and tertiary structures of apo- and holoneocarzinostatin have been investigated by x-ray crystallography and spectroscopic methods. This chapter reviews the determination process of the three-dimensional structure of neocarzinostatin by the combined use of homonuclear two-dimensional nuclear magnetic resonance (NMR) and distance geometry calculations, which has been dramatically improved in the past decade. The protein moiety consists mainly of antiparallel β-sheets forming a β-barrel structure and a backbone sheet. The chromophore molecule is tightly enwrapped by the barrel and thus shielded from exposure to the solvent. The proteinchromophore interaction mechanism is also discussed in the context of release of the active chromophore and the activation mechanism of holo-neocarzinostatin.

Key words

Holo-neocarzinostatin Apo-neocarzinostatin Homonuclear two-dimensional NMR NOE NOESY Distance geometry calculation Restrained molecular dynamics calculation 
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References

  1. 1.
    Wüthrich K (1986) NMR of proteins and nucleic acids. Wiley, New YorkGoogle Scholar
  2. 2.
    Kessler H, Gehrke M, Griesinger C (1988) Two-dimensional NMR spectroscopy: background and overview of the experiments. Angew Chem Int Ed Engl 27: 490–536CrossRefGoogle Scholar
  3. 3.
    Ernst RR, Bodenhausen G, Wokaun A (1986) Principles of nuclear magnetic resonance in one and two dimensions. Clarendon, OxfordGoogle Scholar
  4. 4.
    Clore GM, Gronenborn AM (1991) Structures of larger proteins in solution: three-and four-dimensional heteronuclear NMR spectroscopy. Science 252: 1390–1399PubMedCrossRefGoogle Scholar
  5. 5.
    Wüthrich K (1989) Protein structure determination in solution by nuclear magnetic resonance spectroscopy. Science 243: 45PubMedCrossRefGoogle Scholar
  6. 6.
    Edison AS, Abildgaard F, Westler WM, Mooberry ES, Markley JL (1994) Practical introduction to theory and implementation of multinuclear, multidimensional nuclear magnetic resonance experiments. Methods in Enzymology, vol 239. Academic. New York, pp 3–79Google Scholar
  7. 7.
    Takeshita J, Maeda H, Koike K (1980) Subcellular action of neocarzinostatin. J Biochem (Tokyo) 88: 1071–1080Google Scholar
  8. 8.
    Edo K, Mizugaki M, Koide Y, Seto H, Furihata K, Otake N, Ishida N (1985) The structure of neocarzinostatin chromophore possessing a novel bicyclo[7,3,0] dodecadiyne system. Tetrahedron Lett 26: 331–334CrossRefGoogle Scholar
  9. 9.
    Myers AG (1987) Proposed structure of the neocarzinostatin chromophore-methyl thioglycolate adduct; a mechanism for the nucleophilic activation of neocarzinostatin. Tetrahedron Lett 28: 4493–4496CrossRefGoogle Scholar
  10. 10.
    Kappen LS, Goldberg IH (1989) Identification of 2-deoxyribonolactone at the site of neocarzinostatin-induced cytosine release in the sequence d(AGC). Biochemistry 28: 1027–1032PubMedCrossRefGoogle Scholar
  11. 11.
    Kappen LS, Goldberg IH (1992) Mismatch-induced switch of neocarzinostatin attack sites in the DNA minor groove, Biochemistry 31: 9081–9089PubMedCrossRefGoogle Scholar
  12. 12.
    Meshchwitz SM, Schultz RG, Ashley GW, Goldberg IH (1992) Selective abstraction of 2H from C-l’ of the C residue in AGC. ICT by the radical center at C-2 of activated neocarzinostatin chromophore: structure of the drug/DNA complex responsible for bistranded lesion formation. Biochemistry 31: 9117–9121CrossRefGoogle Scholar
  13. 13.
    Ishida N, Miyazaki K, Kumagai K, Rikimaru M (1965) Neocarzinostatin, and antitumor antibiotic of high molecular weight. Isolation, physicochemical properties and biological activities. J Antibiot (Tokyo) 18: 68–76Google Scholar
  14. 14.
    Koide Y, Ishii F, Hasuda K, Koyama Y, Edo K, Katamine K, Kitame F, Isida N (1980) Isolation of a nonprotein component and a protein component from neocarzinostatin ( NCS) and their biological activities. J Antibiot 33: 342–346Google Scholar
  15. 15.
    Goldberg IH, Hatayama T, Kappen LS, Napier MA, Povirk LF (1981) Molecular actions and targets for cancer chemotherapeutic agents. Academic, New York, pp 163–191Google Scholar
  16. 16.
    Adjadj E. Mispelter J, Quiniou E, Dimicoli JL, Favaudon V, Lhoste JM (1990) Proton NMR studies of apo-neocarzinostatin from Streptomyces carzinostnticus, sequence-specific assignment and secondary structure. Eur J Biochem 190: 263–271Google Scholar
  17. 17.
    Remerowski ML, Glaser SJ, Sicker LC, Samy TSA, Drobny GP (1990) Sequential ‘H-NMR assignments and secondary structure of aponeocarzinostatin in solution. Biochemisty 29: 8401–8409CrossRefGoogle Scholar
  18. 18.
    Gao X, Burkhart W (1991) Two-and three-dimensional proton NMR studies of aponeocarzinostatin. Biochemistry 30: 7730–7739PubMedCrossRefGoogle Scholar
  19. 19.
    Sicker LC, Jensen LH, Samy TSA (1976) Neocarzinostatin: an antitumor protein. A preliminary x-ray diffraction study. Biochem Biophys Res Commun 68: 358–362Google Scholar
  20. 20.
    Teplyakov A, Obmolova G, Wilson K, Kuromizu K (1993) Crystal structure of aponeocarzinostatin at 0.15-nm resolution. Eur J Biochem 213: 737–741PubMedCrossRefGoogle Scholar
  21. 21.
    Takashima H, Amiya S, Kobayashi Y (1991) Neocarzinostatin: interaction between the antitumor-active chromophore and the carrier protein. J Biochem (Tokyo) 109: 807–810Google Scholar
  22. 22.
    Tanaka T, Hirama M, Ueno M, Imajo M, Ishiguro M, Mizugaki M, Edo K, Komatsu H (1991) Proton NMR studies on NCS-chrom binding structure in neocarzinostatin complex. Tetrahedron Lett 32: 3175–3178CrossRefGoogle Scholar
  23. 23.
    Kim KH, Kwon BM, Myers AG, Rees DC (1993) Crystal structure of neocarzinostatin, an antitumor protein-chromophore complex. Science 262: 1042–1046PubMedCrossRefGoogle Scholar
  24. 24.
    Lefevre C, Adjadj E, Quiniou E, Mispelter J (1994) Assignment of the protonated “C resonances of apo-neocarzinostatin by 2D heteronuclear NMR spectroscopy at natural abundance. J Biomolecular NMR 4: 689–702CrossRefGoogle Scholar
  25. 25.
    Mispelter J, Lefevre C, Adjadj E, Quiniou E, Favaudon V (1995) Internal motions of apo-neocarzinostatin as studied by “C-NMR methine relaxation at natural abundance, J Biocool NMR 5: 233–244Google Scholar
  26. 26.
    Constantine KL, Colson KL, Wittekind M, Friedrichs MS, Zein N, Tuttle J, Langley DR, Leet JE, Schroeder DR, Lam KS, Tarmer II BT, Metzler WJ, Bruccoleri RE, Mueller L (1994) Sequential ‘H, ’ 5 C, and “N NMR assignments and solution conformation of apokedarcidin. Biochemistry 33: 11438–11452PubMedCrossRefGoogle Scholar
  27. 27.
    Nagayama K. Kumar A, Wüthrich K, Ernst RR (1980) Experimental techniques of two-dimensional correlated spectroscopy. J Magn Reson 40: 321–334Google Scholar
  28. 28.
    Braunschweiler L, Ernst RR (1983) Coherence transfer by isotropic mixing: application to proton correlation spectroscopy. J Magn Reson 53: 521–528Google Scholar
  29. 29.
    Bax A, Davis DG (1985) MLEV-17-based two-dimensional homonuclear magnetization transfer spectroscopy. J Magn Reson 65: 355–360Google Scholar
  30. 30.
    Kumar A, Ernst RR, Wüthrich K (1980) A two-dimensional nuclear Overhauser enhancement (2D NOE) experiment for the elucidation of complete proton-proton cross-relaxation networks in biological macromolecules. Biochem Biophys Res Commun 95: 1–6PubMedCrossRefGoogle Scholar
  31. 31.
    Macura S, Wüthrich K, Ernst RR (1982) Separation and suppression of coherent transfer effects in two-dimensional NOE and chemical exchange spectroscopy. J Magn Reson 46: 269–282Google Scholar
  32. 32.
    Keepers JW, James TL (1984) A theoretical study of distance determinations from NMR. Two-dimensional nuclear Overhauser effect spectra. J Magn Reson 57: 404–426Google Scholar
  33. 33.
    Hirayama K, Ando T, Takahashi R, Murai A (1986) A revised primary structure of neocarzinostatin using the combination of endopeptidase, carboxypeptidase Y and fast-atom-bombardment mass spectrometry. Bull Chem Soc Jpn 59: 1371–1378CrossRefGoogle Scholar
  34. 34.
    Kuromizu K, Tsunasawa S, Maeda H, Abe O. Sakiyama F (1986) Reexamination of the primary structure of an antitumor protein, neocarzinostatin. Arch Biochem Biophys 246: 199–205CrossRefGoogle Scholar
  35. 35.
    Sakata N, Minamitani S, Kanebe T, Fiori M, Hamada M, Edo K (1993) The amino acid sequence of neocarzinostatin apoprotein deduced from the base sequence of the gene. Biol Pharm Bull 16: 26–28PubMedCrossRefGoogle Scholar
  36. 36.
    Roey PV, Beerman TA (1989) Crystal structure analysis of auromomycin apoprotein (macromomycin) shows importance of protein side chains to chromophore binding selectivity. Proc Natl Acad Sci USA 86: 6587–6591PubMedCrossRefGoogle Scholar
  37. 37.
    Braun W. Go N (1985) Calculation of protein conformations by proton-proton distance constraints. A new efficient algorithm. J Mol Biol 186: 611–626Google Scholar
  38. 38.
    Braun W (1987) Distance geometry and related methods for protein structure determination from NMR data. Q Rev Biophys 19: 115–127PubMedCrossRefGoogle Scholar
  39. 39.
    Kobayashi Y, Ohkubo T, Kyogoku Y, Nishiuchi Y, Sakakibara S, Braun W, Go N (1989) Solution conformation of conotoxin GI determined by ‘H nuclear magnetic resonance spectroscopy and distance geometry calculations. Biochemistry 28: 4853–4860PubMedCrossRefGoogle Scholar
  40. 40.
    Guntert P, Braun W, Wüthrich K (1991) Efficient computation of three-dimensional protein structures in solution from nuclear magnetic resonance data using the program DIANA and the supporting programs CALIGA, HABAS and GLOMSA. J Mol Biol 217: 517–526Google Scholar
  41. 41.
    Guntert P, Wüthrich K (1991) Improved efficiency of protein structure calculations from NMR data using the program DIANA with redundant dihedral angle constraints [REDAC]. J Biomol NMR 1: 447–460PubMedCrossRefGoogle Scholar
  42. 42.
    Borgias BA, James TL (1988) COMATOSE, a method for constrained refinement of macromolecular structure based on two-dimensional nuclear Overhauser effect spectra. J Magn Reson 79: 493–512Google Scholar
  43. 43.
    Andersen NH, Lai XN, Hammen PK, Marschner TM (1990) Computer-aided conformational analysis based on NOESY signal intensities. Basic Life Sci 56: 95–134PubMedGoogle Scholar
  44. 44.
    Tanaka T, Hirama M, Fujita K, lmajo S, Ishiguro M (1993) Solution structure of the antitumor antibiotic neocarzinostatin, a chromophorc-protein complex. J Chem Soc Chem Commun 15: 1205–1207CrossRefGoogle Scholar
  45. 45.
    Mohanty S, Sicker LC, Drobny OP (1994) Sequential ‘H-NMR assignment of the complex of aponeocarzinostatin with ethidium bromide and investigation of protein-drug interactions in NCS-chrom binding site. Biochemistry 33: 10579–10590PubMedCrossRefGoogle Scholar
  46. 46.
    Ishiguro M, Imajo S, Hirama M (1991) Modeling study of the structure of the macro-molecular antitumor antibiotic neocarzinostatin. Origin of the stabilization of NCSchrom. J Med Chem 34: 2366–2373Google Scholar
  47. 47.
    Imajo S, Ishiguro M, Tanaka “l’, Hirama M, Teplyakov A (1995) On the conformation of Phe78 of a chromoprotein antibiotic, neocarzinostatin. Bioorg Med Chem 3: 429–436Google Scholar

Copyright information

© Springer Japan 1997

Authors and Affiliations

  • Hiroyuki Takashima
    • 1
  • Yuji Kobayashi
    • 2
  1. 1.International Research LaboratoriesCiba-Geigy Japan, Ltd.Takarazuka, Hyogo, 655Japan
  2. 2.Faculty of Pharmaceutical SciencesOsaka UniversitySuita, Osaka, 565Japan

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