STRUCTURAL STUDIES IN ENZYME
MECHANISMS AND IN CHEMICAL CARCINOGENESIS AND MUTAGENESIS
JENNY PICKWORTH GLUSKER, D.Phil., D.Sc., Senior Member; Adjunct
Professor of Biochemistry and Biophysics, and Member of Biophysics Graduate
Group, University of Pennsylvania
The stereochemistry of molecular interactions is studied in our laboratory by
means of X-ray diffraction studies of crystalline materials. Our major
interests are in chemical carcinogenesis and mutagenesis, the mechanisms of
enzyme reactions and the roles of metal ions in biological systems.
When a polycyclic aromatic hydrocarbon (PAH) produces a carcinogenic lesion, it has first been activated to a molecular species that can interact with informational macromolecules in the cell. A major mode of activation is the formation of a diol epoxide that then can alkylate DNA. The carcinogenic potential of a PAH is greatly increased if the PAH contains a bay-region methyl group. It has generally been hypothesized that this is because the bay-region methyl group, by virtue of its bulk, causes some significant conformational rigidity in the diol epoxide and/or in the DNA adduct that is formed. The steric hindrance, then, will favor a conformation that facilitates the progression of the carcinogenic process.
Some details of the effects of a bay-region methyl group are found in the crystal structure of the anti-diol epoxide of 5,6-dimethylchrysene, the first experimentally based description of the steric interactions between the bay region methyl group and the ring that bears the diol and epoxide groups. A view of the molecule is shown in Figure 1, together with its adduct with methanol, previously studied in this laboratory. Note that the general orientation of the aromatic ring system is similar in both structures.
ENZYME STRUCTURE AND FUNCTION. SHIMONI-LIVNY, CARRELL, KATZ, WAGNER, AFSHAR, GLUSKER, in collaboration with JAFFE§We are continuing our study of the enzymes, D-xylose isomerase and porphobilinogen synthase, and are now using Dxylose isomerase as a tool for studying its metal binding capabilities and their effects on the catalytic mechanism of this enzyme. Several different metal ions have been added to the apoenzyme and the corresponding crystal structures have been elucidated by Xray diffraction methods. In addition, the
effects of these same cations on the catalytic activity of the enzyme are being studied in an effort to improve our understanding of its mechanism of action.
The enzyme, porphobilinogen synthase, catalyzes the condensation of two identical substrate molecules (5-aminolevulinic acid) in an asymmetric manner to form porphobilinogen. Diffraction data have been obtained for the enzyme from several sources, including Escherichia coli and a bovine liver.
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FIGURE 1. Views of (a) 5,6-dimethylchrysene-1,2-diol-3,4-epoxide with the epoxide and hydroxyl groups indicated by filled bonds and (b) its adduct with methanol (Afshar et al., Carcinogenesis 17:2507, 1994). The view is onto the plane of the epoxide group. |
METAL IONS IN PROTEINS. KATZ,
SHIMONI-LIVNY, WAGNER, BOCK, GEORGE, in collaboration with
TRACHTMANb (2, 12, 13, 14)
The binding of divalent metal ions to proteins is being studied by analyzing of crystal structures containing a selected cation. The nature of ligands in crystal structures of small carbon-containing molecules and in biological macromolecules provides the basis of our analyses. The energetic consequences of changing the coordination environment of the metal ion are determined by ab initio molecular orbital calculations. Our analyses to date involve magnesium, manganese, calcium, zinc and lead. It was found that, if a metal ion-bound water molecule is replaced by a carboxylate group, the ability of the metal ion to polarize remaining water molecules is decreased.
INTERMOLECULAR INTERACTIONS. KATZ, SHIMONI-LIVNY, CARRELL, GLUSKER, in collaboration with CHANDRAKALA,c SAILAJA,c PODILE,c THALLADI,c BRASSELET,c WEISS,c BLAESER,c BOESE,c ZYSS,c NANGIA,c DESIRAJUc (3-5, 15)Our collaboration continues with Professor Gautam Desiraju in his studies of intermolecular interactions and the packing motifs normally found in the crystalline state. One aim is the design of compounds with potential as octupolar nonlinear optical (NLO) materials. The crystal structures and NLO properties of the 2,4,6-triaryloxy-1,3,5-triazines have been reported. These compounds consistently form quasi-trigonal or trigonal networks that are two-dimensionally noncentrosymmetric. The structures we studied were found to be stabilized by weak intermolecular interactions such as herringbone, p· · · p interactions, and C-H· · · O and CH· · · N hydrogen bonding. Other compounds studied include some aza- b-lactams and some isocyanurates. Analyses of packing motifs are currently in progress with the aim of understanding their significance in controlling modes and relative orientations in molecular packing and intermolecular interactions.
ADENOSYLCOBALAMIN CATALYZED REACTIONS. WAGNER, AFSHAR, CARRELL, GEORGE, BOCK, GLUSKER, in collaboration with ENGLERT,d HOGENKAMPe (6)The crystal structure of difluoromethylcobalamin (CF2Cbl), a vitamin B12 analogue with CHF2 replacing CN, has been determined. The packing motif of CF2Cbl is very similar to that described for wet vitamin B12--a distorted hexagonal close packing of the cobalamin with channels of water running parallel to the crystallographic c axis through the crystal at x=1/4, y=0, and x=3/4, y=1/2. An analysis of interactions involving water molecules and side-chain amide groups revealed a discontinuity between oxygen-oxygen distances, which are found to be less than 2.8 Å, and oxygen-nitrogen distances, which are found to be greater than 2.8 Å); this provides a useful criterion for distinguishing between oxygen and nitrogen atoms in the terminal amide groups of B12 structures. A superposition of the crystal structures of vitamin B12 and CF2Cbl shows that in CF2Cbl, the c side chain of ring B takes on a conformation that brings its terminal amide group near to the CHF2 group. As shown in Figure 2, this results in both a relatively short contact (3.11 Å) between the fluorine and the amide oxygen atom, O39, and a weak C1F-H1F· · · O39 interaction.
Our studies of reaction enthalpies and activation energies for the adenosylcobalamin-utilizing enzymes, diol dehydrase, and ethanolamine ammonia-lyase, continue.
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FIGURE 2. View of the difluoro analogy of methylcobalamin showing the orientation and interactions of the -CHF2 group (filled bonds). |
CRYSTALLOGRAPHIC METHODS. CARRELL, GLUSKER
(7-11)
An overall survey of methods of teaching crystallography has been written for the 50th anniversary of Acta Crystallographica. In addition, a review in the morphological studies of macromolecular crystals has been written. I sadly note the death of my textbook co-author, Ken Trueblood.
PUBLICATIONS1. AFSHAR, C.E., KATZ, A.K., CARRELL, H.L., AMIN, S., DESAI, D., GLUSKER, J.P. Three-dimensional structure of 5,6-dimethylchrysene-1,2-diol-3-4-epoxide, a diol epoxide with a bay-region methyl group. Carcinogenesis (in press).
2. KATZ, A.K., GLUSKER, J.P. Roles of zinc and magnesium ions in enzymes. In Advances in Molecular Structure Research, edited by M. Hargittai and I. Hargittai. JAI Press, Stamford, CT, Vol. 5, pp. 227-279, 1998.
3. CHANDRAKALA, P.S., KATZ, A.K., CARRELL, H.L., SAILAJA, P.R., PODILE, A.R., NANGIA, A., DESIRAJU, G.R. Synthesis, X-ray crystal structures and biological evaluation of some mono- and bi-cyclic 1,3-diazetidin-2-ones: non-natural b-lactam analogs. J. Chem. Soc., Perkin Trans. 1:2597-2608, 1998.
4. THALLADI, V.R., BRASSELET, S., WEISS, H.-C., BLAESER, D., KATZ, A.K., CARRELL, H.L., BOESE, R., ZYSS, J., NANGIA, A., DESIRAJU, G.R. Crystal engineering of some 2,4,6-triaryloxy-1,3,5-triazines: octupolar nonlinear materials. J. Am. Chem. Soc. 120:2563-2577, 1998.
5. THALLADI, V.R., KATZ, A.K., CARRELL, H.L., NANGIA, A., DESIRAJU, G.R. Trimethyl isocyanurate and triethyl isocyanurate. Acta Crystallogr. C54:86-89, 1998.
6. WAGNER, T., AFSHAR, C.E., CARRELL, H.L., GLUSKER, J.P., ENGLERT, U., HOGENKAMP, H.P.C. Difluoromethylcobalamin: structural aspects of an old tree with a new branch. Inorg. Chem. (in press).
7. GLUSKER, J.P. The teaching of crystallography: a historical survey. Acta Cryst. A54:707-715, 1998.
8. PARTHASARATHY, S., GLUSKER, J.P. (editors). Aspects of Crystallography in Molecular Biology. New Age International, New Delhi, India, 1997.
9. CARRELL, H.L., GLUSKER, J.P. Crystal morphology, optical properties of crystals and crystal mounting. International Tables for Crystallography, edited by M.G. Rossmann and E. Arnold. Vol. F (in press).
10. GLUSKER, J.P. Kenneth Nyitray Trueblood 1920-1998. Obituary. Acta Cryst. B54:505-506, 1998.
11. EISENBERG, D., GLUSKER, J.P., SPARKS, R. Kenneth Nyitray Trueblood. Obituary. Physics Today, pp. 88-89, September 1998.
Papers in press at time of previous report:
12. GLUSKER, J.P. The binding of ions to proteins. In Protein: A Comprehensive Treatise, edited G. Allen. JAI Press, Stamford, CT, Vol. 2, Ch. 4, pp. 99-152, 1999.
13. TRACHTMAN, M., MARKHAM, G.D., GLUSKER, J.P., GEORGE, P., BOCK, C.W. Interactions of metal ions with water: ab initio orbital studies of structure, bonding enthalpies, vibrational frequencies and charge distributions. Part 1. Monohydrates. Inorganic Chem. 37:4421-4431, 1998.
14. KATZ, A.K., GLUSKER, J.P., MARKHAM, G.D., BOCK, C.W. Deprotonation of water in the presence of carboxylate and magnesium ions. J. Phys. Chem. B102:6342-6350, 1998.
15. GLUSKER, J.P. Directional aspects of intermolecular interactions. In Topics in Current Chemistry, Design of Organic Solids, edited by E. Weber. Springer Verlag, Berlin, Heidelberg, Vol. 198, pp. 1-56, 1998.
KAUFMAN KATZ, A., CARRELL, H.L., GLUSKER, J.P. Dibenzo[a,l]pyrene (dibenzo[def,p]chrysene): fjord-region distortions. Carcinogenesis 19:1641-1648, 1998.
GLUSKER, J.P., CARRELL, H.L., KATZ, A.K., AFSHAR, C.E. Structural studies of PAHS and their metabolites. Polycyclic Aromatic Compounds (in press).
§ Fox Chase researcher
a S. Amin, D. Desai: American Health Foundation, 1 Dana Road, Valhalla, NY 10595
b M. Trachtman: Department of Chemistry, Philadelphia College of Textiles and Science, Philadelphia, PA 19144
c P.S. Chandrakala, P.R. Sailaja, A.R. Podile, V.R. Thalladi, S. Brasselet, H-C. Weiss, D. Blaeser, R. Boese, J. Zyss, A.R. Nangia, G.R. Desiraju: School of Chemistry, University of Hyderabad, Hyderabad, 500046, India
d U. Englert: Institut für Anorganische Chemie, RWTH Aachen, Prof.-Pirlet-Str. 1, 52074 Aachen, Germany
e H.P.C. Hogenkamp: Department of Biochemistry, University of Minnesota, Minneapolis, MN 55455
Illustrations or unpublished data in these reports should not be used without permission of the author.
 Fox Chase Cancer Center |
Scientific Report 1998 |
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