Student Projects for UMass Chemistry 423 Spring 2012-2

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Cisplatin-DNA complex- 1a84

by Greg Keohane, Nicole Hofstetter, Gina Lein, Louis Pires, Student Projects for UMass Chemistry 423 Spring 2012

Introduction

Cisplatin

The figure to the right shows bound to a 12 base pair double stranded DNA. Cisplatin, cis-PtCl2(NH3)2, is an “alkylating” chemotherapy drug, administered intravenously, used in the treatment of various types of cancer. [1]

There are three fundamental components in the mechanism of cisplatin – cisplatin, DNA, and HMG-protein. Cisplatin makes contact with the cell membrane and enters the cell through active transport, but some molecules are passively diffused. This platinum-based drug acts in vivo by to two consecutive adjacent guanine bases in DNA leading to the loss of its chlorine atoms for the nitrogen on the guanine; this occurs to better balance the platinum charge.[2] The binding of cisplatin creates a 49 bend with an overall helix bend of 78, which is crucial to cisplatin’s role as an anticancer drug.[5] The bend in the , as seen in pdb 1ckt, allows for HMG-protein to bind to the DNA, and when bound it inserts a wedge like phenol group of phenylalanine into the widened minor grove. HMG-proteins, high mobility group-proteins, are found everywhere and regulate transcription, replication, recombination and repair, and once bound to the DNA it de-stacks the nucleotide base pairs , which in turn kinks the already mutated DNA. With the HMG-protein bound to the DNA, the cell cannot properly repair the DNA, leading to apoptosis.[3]

This link shows a video on the mechanism of cisplatin.[1]

Unfortunately, there is not yet a definitive way to regulate which cells are affected by cisplatin, so the cytotoxic effects damage normal cells as well, in particular rapidly dividing cells such as those found in the gastrointestinal tract, bone marrow, testicles, ovaries, and hair growth. It is the foundation to many combination treatments for cancers, but not all cancers are effected by cisplatin, the majority of patients using cisplatin will relapse with platinum resistant diseases. Another way cisplatin can be in effective, is when the cancers gets too old; when a tumor starts out it divides more frequently and this is when it is effected. This same logic goes for solid tumors, other treatments are needed for these types of issues.


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Overall Structure

This conformation shown above has the widening and flattening of the minor groove of the DNA molecule which resembles A-DNA not found in B DNA. This change is due to the guanine bases that cisplatin interacts with. They compact the major groove and unwind the DNA.

The original view shows a double stranded DNA helix and the cisplatin ligand. It is a "duplex dodecamer" d(CCTCTG*G*TCTCCGGAGACCAGAGG), and the asterisks are denoting which base pairs the Ciplatin binds to. This molecule is in its which means it stll has the right handed helix, but the widened minor groove distorts its structure. To prove this, the animation shows the distance between intrastrand phosphate groups has been changed due to the insertion of the Cisplatin. Based on the chart found in (Forms of DNA), the distance should changed from 7 [Å] to 5.9 [Å]. It is much more rare than the common B-DNA[2] [4].


. The cisplatin ligand is a cis-diammineplatinum molecule, which is a platinum atom attached to two N7 nitrogen atoms, each apart of a guanine bases, and two NH3 molecules attached to the other side. They attach to the 6 and 7 guanine bases which links the two bases together and alters the bend in the helix by 49 degrees.The guanine still pair with the 18 and 19 cytosine bases.[5]

Normally, the Cisplatin molecule has two Cl atoms attached to Pt's last two electrons [PtCl2(NH3)2]. The Cl atoms can be displaced by "aquation" to form [PtCl(H2O)(NH3)2]+. The resulting ligand can be linked to bases now and guanine is the preferred choice. It can then cross-link as in the Cisplatin molecules shown above to form [Pt(guanine-DNA)2(NH3)2]+.[6] The N-Pt-N angles, two N7 nitrogens from the guanine base and two NH3 attached directly as part of the Cisplatin ligand, are planar and at 90 degrees, as well as each being a distance of 2.05 [Å] from the PT atom. Another interesting feature about the platinated lesion is in a 5 base section between C4-G21 and T8-A17. The minor groove of the molecule is roughly 9-12 [Å] with a depth range of roughly .4-2.5 [Å] which is much shorter than the B form with a depth of 6.5 [Å][7].






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Binding Interactions

Cisplatin

As described above, the cisplatin ligand binds to the N7 atoms of the adjacent G6 and G7 guanine bases in a strand of DNA. The green screen shows the platinum atom in pink, which is bound to the two N7 atoms of gaunine labeled in green. The N7 atoms are bound to the platinum atom in the ligand, creating a bend in the helix towards the guanine bases of 49 degrees and a total bend in the DNA of 79 degrees[8]. The guanine bases are favored over the adenine because of hydrogen bonding between the amine-hydrogens of the cisplatin and the O=C6 moiety of guanine[9]. The is shown in this green screen, between the green labeled ammine hydrogens and the oxygen atom labeled in pink. The resulting platination also causes the duplex to unwind by approximately 25 degrees at the site of platination from the base pair T8-A17 to T5-A20. The screen shows the thymine molecules in pink and the adenine ones in green, showing the opening of the helix. These distortions in the duplex allow the minor groove opposite the platinum to be opened to 9.0-12 angstroms, making it shallow and wide.

HMG-domain proteins (High-mobility group) bind to recognition sequences found in the minor groove. The bend in DNA caused by cisplatin leaves the minor groove more vulnerable and open for recognition by HMG-domain proteins. Since the expression of these proteins are correlated to tumor cells, the recognition of them by cisplatin-bound DNA could lead to a therapy of cancerous tumors[10].








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Additional Features

The width of the minor groove of the cisplatin-DNA complex can be compared to the minor groove width of HMG protein-DNA complexes. The similar widths are evidence that Cisplatin is affiliated with HMG proteins.

Structural studies of Cisplatin-modified DNA are underway in hope to find a significant correlation between Cisplatin distorted DNA and its ability to bind to high mobility group proteins (HMG proteins). HMG proteins are responsible for many actions within the cell such as transcription, replication and DNA repair. Studies show that over/under expression of these proteins may be the cause of tumors. On the other hand, if HMG proteins attach to a Cisplatin modified double helix, then this may prevent the excision of the helix to be repaired. Resulting in DNA destruction.

HMG proteins bind to the minor groove of the DNA duplex where their recognition sequences are located. Cisplatin is known to cause the bending of DNA helix as well as the extension of the minor groove width. This opening of the minor groove allows HMG domain proteins to attached to their recognition sequences within the minor groove DNA base pairs

This deformation of the DNA duplex with cisplatin forms important complexes with HMG proteins, such as LEF-1 and hSRY. Binding of these proteins to the already damaged DNA causes further bending. The LEF-1 HMG protein structure was determined by experiment and superimposed over the known cisplatin-DNA structure.[11] The best fit was shown be over the portion of the cisplatin-DNA structure containing the platinated guanonsines of the 1,2 intrastrand cross link, the similarity holds a good overlap of RMSD of 3.2A.

This experiment showed that the distortion caused by the LEF-1 protein is very similar to that caused buy the binding of cisplatin. This comparison of LEF-1 to cisplatin-modified DNA brings structural evidence that cisplatin –modified DNA may signal the recognition of HMG proteins.

Another important example is the HMG1 protein binding to the Cisplatin complex. This HMG1protein is known to bind to the Cisplatin DNA minor groove about the hydrophobic kink created by the distortion. Evidence shows that the phenylalanine residue HMG1 protein is essential for HMG1 interaction with DNA. [12]Substitution experiments of the phenylalanine with alanine showed that HMG1HMG1 binding reduced, therefore HMG1 binding is dependent on the phenylalanine and the hydrophobic notch

Credits

Introduction - Gina Lein

Overall Structure - Greg Keohane

Drug Binding Site - Louis Pires

Additional Features - Nicole Hofstetter

References

  1. adhttp://en.wikipedia.org/wiki/Chemotherapy
  2. David, G.S. The Molecular Perspective: Cisplatin. doi: 10.1634/theoncologist.11-3-316 The Oncologist March 2006 vol. 11 no. 3 316-317.
  3. Gelasco, Andrew. "NMR solution and structure of DNA Dodecamer Duplex Containing cis-Diammaineplatium" Department of Chemistry, MIT:1998
  4. Takahara, P. M., Rosenzweig, A. C., Frederick, C. A., and Lippard, S. J. (1995) Nature 377, 649-652. Takahara, P. M., Frederick, C. A., and Lippard, S. J. (1996) J. Am. Chem. Soc 118, 12309-12321
  5. Fichtinger-Schepman, A. M. J., van der Veer, J. L., den Hartog, J. H. J., Lohman, P. H. M., and Reedijk, J. (1985) Biochemistry 24, 707-713.
  6. http://en.wikipedia.org/wiki/Cisplatin,last accessed 4/8/12.
  7. Andrew Gelasco and Stephen J. Lippard* Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 ReceiVed December 30, 1997; ReVised Manuscript ReceiVed March 27, 1998
  8. Gelasco, Andrew. "NMR solution and structure of DNA Dodecamer Duplex Containing cis-Diammaineplatium" Department of Chemistry, MIT:1998
  9. Baik MH, Friesner RA, Lippard SJ. "Theoretical Study of Cisplatin binding to purine bases: why doe cisplatin prefer guanine over adenine?" J Am Chem Soc, 2003 Nov 19;125(46):14082-92. 1.↑ 2.0 2.1 2.2 2.3 2.4 2.5 Wang G, Vasquez KM. Z-DNA, an active element in the genome. Front Biosci. 2007 May 1;12:4424-38. PMID:17485386
  10. Gelasco, Andrew. "NMR solution and structure of DNA Dodecamer Duplex Containing cis-Diammaineplatium" Department of Chemistry, MIT:1998
  11. Gelasco, Andrew. "NMR solution and structure of DNA Dodecamer Duplex Containing cis-Diammaineplatium" Department of Chemistry, MIT:1998
  12. Love, JJ. "Structural basis for DNA bending by the architectural transcription factor LEF-1." PubMed:1995 http://www.rcsb.org/pdb/explore/explore.do?structureId=2LEF

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