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The MOVE Fire

This is a personal recollection on the Move fire on May 13, 1985 Philadelphia police fired thousands of rounds at the MOVE house, city officials approved dropping an explosive device on the roof, the resulting fire was allowed to burn, 11 people—including five children—died, and 61 homes were destroyed. Philadelphia City Council later called it a “brutal attack carried out by the City of Philadelphia on its own citizens” and acknowledged that no individual faced criminal consequences for the bombing. One timeline correction worth preserving for the BHP record: the major previous MOVE-police confrontation was August 8, 1978, about seven years before the bombing, not a year or two earlier. Officer James Ramp was killed, other police and firefighters were wounded, nine MOVE members were later convicted, and television cameras recorded police beating Delbert Africa during his arrest. The 1985 MOVE Commission later specifically criticized city planners for failing to adequately use lessons from that 1978 confrontation. And that actually strengthens the point you’re making: 1985 did not happen without precedent or institutional memory. There had already been a deadly confrontation with MOVE, years of conflict, negotiations and police involvement before Osage Avenue.

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BLACK FACTS
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The Violence That Helped Spark the NAACP

In August 1908, a white mob attacked Springfield, Illinois’s Black community, destroying homes and businesses and lynching two Black men. National outrage over the violence helped spur the movement that created the NAACP the following year.

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Wikipedia

DNA base flipping

Cartoon of DNA with a base flipped out
The DNA double helix with a cytosine base flipped out 180°

DNA base flipping, or nucleotide flipping, is a mechanism in which a single nucleotide base, or nucleobase, is rotated outside the nucleic acid double helix.[1] This occurs when a nucleic acid-processing enzyme needs access to the base to perform work on it, such as its excision for replacement with another base during DNA repair. It was first observed in 1994 using X-ray crystallography in a methyltransferase enzyme catalyzing methylation of a cytosine base in DNA. Since then, it has been shown to be used by different enzymes in many biological processes such as DNA methylation, various DNA repair mechanisms, and DNA replication. It can also occur in RNA double helices[2] or in the DNA:RNA intermediates formed during RNA transcription.

DNA base flipping occurs by breaking the hydrogen bonds between the bases and unstacking the base from its neighbors. This could occur through an active process, where an enzyme binds to the DNA and then facilitates rotation of the base, or a passive process, where the base rotates out spontaneously, and this state is recognized and bound by an enzyme. It can be detected using X-ray crystallography, NMR spectroscopy, fluorescence spectroscopy, or hybridization probes.

Discovery

[edit]

Base flipping was first observed in 1994 when researchers Klimasauskas, Kumar, Roberts, and Cheng used X-ray crystallography to view an intermediate step in the chemical reaction of a methyltransferase bound to DNA.[3] The methyltransferase they used was the C5-cytosine methyltransferase from Haemophilus haemolyticus (M. HhaI). This enzyme recognizes a specific sequence of the DNA (5'-GCGC-3') and methylates the first cytosine base of the sequence at its C5 location.[3] Upon crystallization of the M. HhaI-DNA complex, they saw the target cytosine base was rotated completely out of the double helix and was positioned in the active site of the M. HhaI. It was held in place by numerous interactions between the M. HhaI and DNA.[3]

The authors theorized that base flipping was a mechanism used by many other enzymes, such as helicases, recombination enzymes, RNA polymerases, DNA polymerases, and Type II topoisomerases.[3] Much research has been done in the years subsequent to this discovery and it has been found that base flipping is a mechanism used in many of the biological processes the authors suggest.[4][5][6]

Mechanism

[edit]
Model of interactions between DNA with flipped base and a methyltransferase
Model of Entamoeba histolytica DNMT2. Demonstrates a base flipped out of the double helix and into the active site of a methyltransferase.

DNA nucleotides are held together with hydrogen bonds, which are relatively weak and can be easily broken. Base flipping occurs on a millisecond timescale[7] by breaking the hydrogen bonds between bases and unstacking the base from its neighbors.[8] The base is rotated out of the double helix by 180 degrees,[9] typically via the major groove,[5] and into the active site of an enzyme. This opening leads to small conformational changes in the DNA backbone[10] which are quickly stabilized by the increased enzyme-DNA interactions.[5] Studies looking at the free-energy profiles of base flipping have shown that the free-energy barrier to flipping can be lowered by 17 kcal/mol for M.HhaI in the closed conformation.[5]

There are two mechanisms of DNA base flipping: active and passive.[11] In the active mechanism, an enzyme binds to the DNA and then actively rotates the base, while in the passive mechanism a damaged base rotates out spontaneously first, then is recognized and bound by the enzyme.[8] Research has demonstrated both mechanisms: uracil-DNA glycosylase follows the passive mechanism[8] and Tn10 transposase follows the active mechanism.[12]

Furthermore, studies have shown that DNA base flipping is used by many different enzymes in a variety biological processes such as DNA methylation, various DNA repair mechanisms, RNA transcription and DNA replication.[4][5][6]

Biological processes

[edit]

DNA modification and repair

[edit]
Model of uracil DNA glycosylase and flipped uracil residue
A uracil residue flipped out of the DNA double helix and into the specificity pocket of Uracil DNA glycosylase

DNA can have mutations that cause a base in the DNA strand to be damaged. To ensure genetic integrity of the DNA, enzymes need to repair any damage. There are many types of DNA repair. Base excision repair utilizes base flipping to flip the damaged base out of the double helix[5] and into the specificity pocket of a glycosylase which hydrolyzes the glycosidic bond and removes the base.[13] DNA glycosylases interact with DNA, flipping bases to determine a mismatch. An example of base excision repair occurs when a cytosine base is deaminated and becomes a uracil base. This causes a U:G mispair which is detected by Uracil DNA glycosylase. The uracil base is flipped out into the glycosylase active pocket where it is removed from the DNA strand.[14] Base flipping is used to repair mutations such as 8-Oxoguanine (oxoG)[15] and thymine dimers created by UV radiation.[13][16]

Replication, transcription and recombination

[edit]

DNA replication and RNA transcription both make use of base flipping.[5] DNA polymerase is an enzyme that carries out replication. It can be thought of as a hand that grips the DNA single strand template.[13] As the template passes across the palm region of the polymerase, the template bases are flipped out of the helix and away from the dNTP binding site.[17] During transcription, RNA polymerase catalyzes RNA synthesis. During the initiation phase, two bases in the -10 element flip out from the helix and into two pockets in RNA polymerase. These new interactions stabilize the -10 element and promote the DNA strands to separate or melt.[13][18]

Base flipping occurs during latter stages of recombination.[19] RecA is a protein that promotes strand invasion[13] during homologous recombination. Base flipping has been proposed as the mechanism by which RecA can enable a single strand to recognize homology in duplex DNA.[20] Other studies indicate that it is also involved in V(D)J Recombination.[21]

DNA methylation

[edit]
DNA methylation illustration
DNA molecule that is methylated on both strands on the center cytosine

DNA methylation is the process in which a methyl group is added to either a cytosine or adenine.[22] This process causes the activation or inactivation of gene expression, thereby resulting in gene regulation in eukaryotic cells. DNA methylation process is also known to be involved in certain types of cancer formation.[23][24][25] In order for this chemical modification to occur, it is necessary that the target base flips out of the DNA double helix to allow the methyltransferases to catalyze the reaction.[5]

Target recognition by restriction endonucleases

[edit]

Restriction endonucleases, also known as restriction enzymes are enzymes that cleave the sugar-phosphate backbone of the DNA at specific nucleotides sequences that are usually four to six nucleotides long.[26] Studies performed by Horton and colleagues have shown that the mechanism by which these enzymes cleave the DNA involves base flipping as well as bending the DNA and the expansion of the minor groove.[27] In 2006, Horton and colleagues, x-ray crystallography evidence was presented showing that the restriction endonuclease HinP1I utilizes base flipping in order to recognize its target sequence. This enzyme is known to cleave the DNA at the palindromic tetranucleotide sequence G↓CGC.

Experimental approaches for detection

[edit]

X-ray crystallography

[edit]
X-ray crystallography workflow
Workflow for solving the structure of a molecule by X-ray crystallography

X-ray crystallography is a technique that measures the angles and intensities of crystalline atoms in order to determine the atomic and molecular structure of the crystal of interest. Crystallographers are then able to produce and three-dimensional picture where the positions of the atoms, chemical bonds as well as other important characteristics can be determined.[citation needed] Klimasaukas and colleagues used this technique to observe the first base flipping phenomenon, in which their experimental procedure involved several steps:[3]

  1. Purification
  2. Crystallization
  3. Data Collection
  4. Structure determination and refinement

During purification, Haemophilus haemolyticus methyltransferase was overexpressed and purified using a high salt back-extraction step to selectively solubilize M.HhaI, followed by fast protein liquid chromatography (FPLC) as done previously by Kumar and colleagues.[28] Authors utilized a Mono-Q anion exchange column to remove the small quantity of proteinaceous materials and unwanted DNA prior to the crystallization step. Once M.HhaI was successfully purified, the sample was then grown using a method that mixes the solution containing the complex at a temperature of 16 °C and the hanging-drop vapor diffusion technique to obtain the crystals. Authors were then able to collect the x-ray data according to a technique used by Cheng and colleagues in 1993.[29] This technique involved the measurement of the diffraction intensities on a FAST detector, where the exposure times for 0.1° rotation were 5 or 10 seconds. For the structure determination and refinement, Klimasaukas and colleagues used the molecular replacement of the refined apo structure described by Cheng and colleagues in 1993[29] where the search models X-PLOR, MERLOT, and TRNSUM were used to solve the rotation and translation functions.[30][31] This part of the study involves the use of a variety of software and computer algorithms to solve the structures and characteristics of the crystal of interest.

NMR spectroscopy

[edit]

NMR spectroscopy is a technique that has been used over the years to study important dynamic aspects of base flipping. This technique allows researchers to determine the physical and chemical properties of atoms and other molecules by utilizing the magnetic properties of atomic nuclei.[citation needed] In addition, NMR can provide a variety of information including structure, reaction states, chemical environment of the molecules, and dynamics.[32][33] During the DNA base flipping discovery experiment, researchers utilized NMR spectroscopy to investigate the enzyme-induced base flipping of HhaI methyltransferase. In order to accomplish this experiment, two 5-fluorocytosine residues were incorporated into the target and the reference position with the DNA substrate so the 19F chemical shift analysis could be performed. Once the 19F chemical shift analysis was evaluated, it was then concluded that the DNA complexes existed with multiple forms of the target 5-fluorocytosine along the base flipping pathway.[34]

Fluorescence spectroscopy

[edit]

Fluorescence spectroscopy is a technique that is used to assay a sample using a fluorescent probe. DNA nucleotides themselves are not good candidates for this technique because they do not readily re-emit light upon light excitation.[34] A fluorescent marker is needed to detect base flipping. 2-Aminopurine is a base that is structurally similar to adenine, but is very fluorescent when flipped out from the DNA duplex.[35] It is commonly used to detect base flipping and has an excitation at 305‑320 nm and emission at 370 nm so that it well separated from the excitations of proteins and DNA. Other fluorescent probes used to study DNA base flipping are 6MAP (4‑amino‑6‑methyl‑7(8H)‑pteridone)[36] and Pyrrolo‑C (3-[β-D-2-ribofuranosyl]-6-methylpyrrolo[2,3-d]pyrimidin-2(3H)-one).[37][38] Time-resolved fluorescence spectroscopy is also employed to provide a more detailed picture of the extent of base flipping as well as the conformational dynamics occurring during base flipping.[39]

Hybridization probing

[edit]

Hybridization probes can be used to detect base flipping. This technique uses a molecule that has a complementary sequence to the sequence you would like to detect such that it binds to a single-strand of the DNA or RNA. Several hybridization probes have been used to detect base flipping. Potassium permanganate is used to detect thymine residues that have been flipped out by cytosine-C5 and adenine-N6 methyltransferases.[40] Chloroacetaldehyde is used to detect cytosine residues flipped out by the HhaI DNA cytosine-5 methyltransferase (M. HhaI).[41]

DNA showing a hybridization probe
A hybridization probe is added to a DNA molecule.

See also

[edit]

References

[edit]
  1. ^ Roberts, Richard J.; Cheng, Xiaodong (June 1998). "BASE FLIPPING". Annual Review of Biochemistry. 67 (1): 181–198. doi:10.1146/annurev.biochem.67.1.181. PMID 9759487.
  2. ^ Reiter, Nicholas J.; Blad, Heike; Abildgaard, Frits; Butcher, Samuel E. (1 November 2004). "Dynamics in the U6 RNA Intramolecular Stem−Loop: A Base Flipping Conformational Change". Biochemistry. 43 (43): 13739–13747. doi:10.1021/bi048815y. PMID 15504036. S2CID 25391616.
  3. ^ a b c d e Klimasauskas, Saulius; Kumar, Sanjay; Roberts, Richard J.; Cheng, Xiaodong (January 1994). "Hhal methyltransferase flips its target base out of the DNA helix". Cell. 76 (2): 357–369. doi:10.1016/0092-8674(94)90342-5. PMID 8293469. S2CID 23161543.
  4. ^ a b Brown, Tom. "Nucleic Acids Book". ATDBio. Retrieved 26 February 2014.
  5. ^ a b c d e f g h Huang, Niu; Banavali, Nilesh K.; MacKerell, Alexander D. (7 January 2003). "Protein-facilitated base flipping in DNA by cytosine-5-methyltransferase". Proceedings of the National Academy of Sciences. 100 (1): 68–73. Bibcode:2003PNAS..100...68H. doi:10.1073/pnas.0135427100. PMC 140885. PMID 12506195.
  6. ^ a b Grubmüller, Helmut. "DNA Base Flipping". The Max Planck Institute for Biophysical Chemistry. Archived from the original on 4 February 2017. Retrieved 26 February 2014.
  7. ^ Bouvier, Benjamin; Grubmüller, Helmut (August 2007). "A Molecular Dynamics Study of Slow Base Flipping in DNA using Conformational Flooding". Biophysical Journal. 93 (3): 770–786. Bibcode:2007BpJ....93..770B. doi:10.1529/biophysj.106.091751. PMC 1913169. PMID 17496048.
  8. ^ a b c Larivière, Laurent; Moréra, Solange (August 2004). "Structural Evidence of a Passive Base-flipping Mechanism for β-Glucosyltransferase". Journal of Biological Chemistry. 279 (33): 34715–34720. doi:10.1074/jbc.M404394200. PMID 15178685.
  9. ^ Grosjean, Henri, ed. (2009). DNA and RNA modification enzymes: structure, mechanism, function and evolution. Molecular biology intelligence unit. Austin, Tex.: Landes Bioscience. ISBN 978-1-58706-329-9. OCLC 316421190. Archived from the original on 2014-04-07. Retrieved 2014-03-10.
  10. ^ Giudice, Emmanuel; Várnai, Péter; Lavery, Richard (1 March 2003). "Base pair opening within B-DNA: free energy pathways for GC and AT pairs from umbrella sampling simulations". Nucleic Acids Research. 31 (5): 1434–1443. doi:10.1093/nar/gkg239. PMC 149832. PMID 12595551.
  11. ^ O'Neil, Lauren L. (2008). Base Flipping: Detection, Structures and Energetics (Thesis). University of Notre Dame. doi:10.7274/9306sx63d8n.
  12. ^ Bischerour, Julien; Chalmers, Ronald (10 July 2009). "Base Flipping in Tn10 Transposition: An Active Flip and Capture Mechanism". PLOS ONE. 4 (7) e6201. Bibcode:2009PLoSO...4.6201B. doi:10.1371/journal.pone.0006201. PMC 2705183. PMID 19593448.
  13. ^ a b c d e Watson, James D.; Baker, Tania A.; Bell, Stephen P.; Gann, Alexander; Levine, Michael; Losick, Richard (2014). Molecular biology of the gene (Seventh ed.). Boston: Pearson. ISBN 978-0-321-76243-6.
  14. ^ Krokan, Hans E; Drabløs, Finn; Slupphaug, Geir (16 December 2002). "Uracil in DNA – occurrence, consequences and repair". Oncogene. 21 (58): 8935–8948. doi:10.1038/sj.onc.1205996. PMID 12483510.
  15. ^ Banerjee, Anirban; Yang, Wei; Karplus, Martin; Verdine, Gregory L. (March 2005). "Structure of a repair enzyme interrogating undamaged DNA elucidates recognition of damaged DNA". Nature. 434 (7033): 612–618. Bibcode:2005Natur.434..612B. doi:10.1038/nature03458. PMID 15800616. S2CID 4426014.
  16. ^ Fuxreiter, Monika; Luo, Ning; Jedlovszky, Pál; Simon, István; Osman, Roman (November 2002). "Role of Base Flipping in Specific Recognition of Damaged DNA by Repair Enzymes". Journal of Molecular Biology. 323 (5): 823–834. doi:10.1016/S0022-2836(02)00999-3. hdl:2437/124488. PMID 12417196.
  17. ^ Patel, Premal H.; Suzuki, Motoshi; Adman, Elinor; Shinkai, Akeo; Loeb, Lawrence A. (May 2001). "Prokaryotic DNA polymerase I: evolution, structure, and "base flipping" mechanism for nucleotide selection". Journal of Molecular Biology. 308 (5): 823–837. doi:10.1006/jmbi.2001.4619. PMID 11352575.
  18. ^ Lim, Heon Man; Lee, Hee Jung; Roy, Siddhartha; Adhya, Sankar (18 December 2001). "A "master" in base unpairing during isomerization of a promoter upon RNA polymerase binding". Proceedings of the National Academy of Sciences. 98 (26): 14849–14852. Bibcode:2001PNAS...9814849L. doi:10.1073/pnas.261517398. PMC 64947. PMID 11734629.
  19. ^ Voloshin, Oleg N.; Camerini-Otero, R.Daniel (September 2004). "Synaptic Complex Revisited". Molecular Cell. 15 (6): 846–847. doi:10.1016/j.molcel.2004.09.010. PMID 15383274.
  20. ^ Folta-Stogniew, Ewa; O'Malley, Shawn; Gupta, Ravindra; Anderson, Karen S.; Radding, Charles M. (September 2004). "Exchange of DNA Base Pairs that Coincides with Recognition of Homology Promoted by E. coli RecA Protein". Molecular Cell. 15 (6): 965–975. doi:10.1016/j.molcel.2004.08.017. PMID 15383285.
  21. ^ Bischerour, Julien; Lu, Catherine; Roth, David B.; Chalmers, Ronald (1 November 2009). "Base Flipping in V(D)J Recombination: Insights into the Mechanism of Hairpin Formation, the 12/23 Rule, and the Coordination of Double-Strand Breaks". Molecular and Cellular Biology. 29 (21): 5889–5899. doi:10.1128/MCB.00187-09. PMC 2772739. PMID 19720743.
  22. ^ Klose, Robert J.; Bird, Adrian P. (February 2006). "Genomic DNA methylation: the mark and its mediators". Trends in Biochemical Sciences. 31 (2): 89–97. doi:10.1016/j.tibs.2005.12.008. ISSN 0968-0004. PMID 16403636.
  23. ^ Nakao, Mitsuyoshi (October 2001). "Epigenetics: interaction of DNA methylation and chromatin". Gene. 278 (1–2): 25–31. doi:10.1016/s0378-1119(01)00721-1. PMID 11707319.
  24. ^ Plass, Christoph; Soloway, Paul D (1 January 2002). "DNA methylation, imprinting and cancer". European Journal of Human Genetics. 10 (1): 6–16. doi:10.1038/sj.ejhg.5200768. PMID 11896451.
  25. ^ Esteller, Manel; Herman, James G. (January 2002). "Cancer as an epigenetic disease: DNA methylation and chromatin alterations in human tumours". The Journal of Pathology. 196 (1): 1–7. doi:10.1002/path.1024. PMID 11748635. S2CID 35380651.
  26. ^ "Biology and Activity of Restriction Endonucleases". Archived from the original on 2014-04-18. Retrieved 2014-04-03.
  27. ^ Horton, John R.; Zhang, Xing; Maunus, Robert; Yang, Zhe; Wilson, Geoffrey; Roberts, Richard; Cheng, Xiaodong (2006). "DNA Nicking by HinP1I Endonuclease: Bending, Base Flipping and Minor Groove Expansion". Nucleic Acids Research. 34 (3): 939–948. doi:10.1093/nar/gkj484. PMC 1363774. PMID 16473850.
  28. ^ Kumar, Sanjay; Cheng, Xiaodong; Pflugrath, James W.; Roberts, Richard J. (15 September 1992). "Purification, crystallization, and preliminary x-ray diffraction analysis of an M.HhaI-AdoMet complex". Biochemistry. 31 (36): 8648–8653. doi:10.1021/bi00151a035. PMID 1390649.
  29. ^ a b Cheng, Xiaodong; Kumar, Sanjay; Posfai, Janos; Pflugrath, James W.; Roberts, Richard J. (July 1993). "Crystal structure of the Hhal DNA methyltransferase complexed with S-adenosyl-l-methionine". Cell. 74 (2): 299–307. doi:10.1016/0092-8674(93)90421-l. PMID 8343957. S2CID 54238106.
  30. ^ Brünger, Axel T. (1992). X-PLOR, Version 3.1: a system for X-ray crystallography and NMR. New Haven: Yale University Press. ISBN 978-0-300-05402-6.
  31. ^ Fitzgerald, P. M. D. (1 June 1988). "MERLOT, an integrated package of computer programs for the determination of crystal structures by molecular replacement". Journal of Applied Crystallography. 21 (3): 273–278. Bibcode:1988JApCr..21..273F. doi:10.1107/s0021889887012299.
  32. ^ Guéron, Maurice; Leroy, Jean-Louis (1995). Studies of base pair kinetics by NMR measurement of proton exchange. Methods in Enzymology. Vol. 261. pp. 383–413. doi:10.1016/s0076-6879(95)61018-9. ISBN 978-0-12-182162-3. PMID 8569504.
  33. ^ Leijon, M.; Graslund, A. (1992). "Effects of sequence and length on imino proton-exchange and basepair opening kinetics in DNA oligonucleotide duplexes". Nucleic Acids Res. 20 (20): 5339–5343. doi:10.1093/nar/20.20.5339. PMC 334339. PMID 1331987.
  34. ^ a b Klimašauskas, Saulius; Liutkevičiūtė, Zita (2009). "Experimental Approaches to Study DNA Base Flipping". In Grosjean, Henri (ed.). DNA and RNA modification enzymes: structure, mechanism, function and evolution. Austin, Texas: Landes Bioscience. ISBN 978-1-58706-329-9.
  35. ^ Holz, B (15 February 1998). "2-Aminopurine as a fluorescent probe for DNA base flipping by methyltransferases". Nucleic Acids Research. 26 (4): 1076–1083. doi:10.1093/nar/26.4.1076. PMC 147370. PMID 9461471.
  36. ^ Yang, Kongsheng; Matsika, Spiridoula; Stanley, Robert J. (1 September 2007). "6MAP, a Fluorescent Adenine Analogue, Is a Probe of Base Flipping by DNA Photolyase". The Journal of Physical Chemistry B. 111 (35): 10615–10625. doi:10.1021/jp071035p. PMID 17696385.
  37. ^ Yang, Kongsheng; Stanley, Robert J. (May 2008). "The Extent of DNA Deformation in DNA Photolyase– Substrate Complexes: A Solution State Fluorescence Study". Photochemistry and Photobiology. 84 (3): 741–749. doi:10.1111/j.1751-1097.2007.00251.x. PMID 18086248. S2CID 44506405.
  38. ^ Berry, David A.; Jung, Kee-Yong; Wise, Dean S.; Sercel, Anthony D.; Pearson, William H.; Mackie, Hugh; Randolph, John B.; Somers, Robert L. (March 2004). "Pyrrolo-dC and pyrrolo-C: fluorescent analogs of cytidine and 2′-deoxycytidine for the study of oligonucleotides". Tetrahedron Letters. 45 (11): 2457–2461. doi:10.1016/j.tetlet.2004.01.108.
  39. ^ Neely, Robert K.; Tamulaitis, Gintautas; Chen, Kai; Kubala, Marta; Siksnys, Virginijus; Jones, Anita C. (November 2009). "Time-resolved fluorescence studies of nucleotide flipping by restriction enzymes". Nucleic Acids Research. 37 (20): 6859–6870. doi:10.1093/nar/gkp688. PMC 2777440. PMID 19740769.
  40. ^ Serva, Saulius; Weinhold, Elmar; Roberts, Richard J.; Klimasauskas, Saulius (1 August 1998). "Chemical display of thymine residues flipped out by DNA methyltransferases". Nucleic Acids Research. 26 (15): 3473–3479. doi:10.1093/nar/26.15.3473. PMC 147733. PMID 9671807.
  41. ^ Daujotyte, D.; Liutkeviciute, Z.; Tamulaitis, G.; Klimasauskas, S. (15 April 2008). "Chemical mapping of cytosines enzymatically flipped out of the DNA helix". Nucleic Acids Research. 36 (10): e57. doi:10.1093/nar/gkn200. PMC 2425465. PMID 18450817.

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Wikipedia

DNA base flipping

DNA base flipping, or nucleotide flipping, is a mechanism in which a single nucleotide base, or nucleobase, is rotated outside the nucleic acid double helix. This occurs when a nucleic acid-processing enzyme needs access to the base to perform work on it, such as its excision for replacement with another base during DNA repair. It was first observed in 1994 using X-ray crystallography in a methyltransferase enzyme catalyzing methylation of a cytosine base in DNA. Since then, it has been shown to be used by different enzymes in many biological processes such as DNA methylation, various DNA repair mechanisms, and DNA replication. It can also occur in RNA double helices or in the DNA:RNA intermediates formed during RNA transcription. DNA base flipping occurs by breaking the hydrogen bonds between the bases and unstacking the base from its neighbors. This could occur through an active process, where an enzyme binds to the DNA and then facilitates rotation of the base, or a passive process, where the base rotates out spontaneously, and this state is recognized and bound by an enzyme. It can be detected using X-ray crystallography, NMR spectroscopy, fluorescence spectroscopy, or hybridization probes.

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Wikipedia

Base pair

A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA and RNA. Dictated by specific hydrogen bonding patterns, "Watson–Crick" (or "Watson–Crick–Franklin") base pairs (guanine–cytosine and adenine–thymine/uracil) allow the DNA helix to maintain a regular helical structure that is subtly dependent on its nucleotide sequence. The complementary nature of this based-paired structure provides a redundant copy of the genetic information encoded within each strand of DNA. The regular structure and data redundancy provided by the DNA double helix make DNA well suited to the storage of genetic information, while base-pairing between DNA and incoming nucleotides provides the mechanism through which DNA polymerase replicates DNA and RNA polymerase transcribes DNA into RNA. Many DNA-binding proteins can recognize specific base-pairing patterns that identify particular regulatory regions of genes. Intramolecular base pairs can occur within single-stranded nucleic acids. This is particularly important in RNA molecules (e.g., transfer RNA), where Watson–Crick base pairs (guanine–cytosine and adenine-uracil) permit the formation of short double-stranded helices, and a wide variety of non–Watson–Crick interactions (e.g., G–U or A–A) allow RNAs to fold into a vast range of specific three-dimensional structures. In addition, base-pairing between transfer RNA (tRNA) and messenger RNA (mRNA) forms the basis for the molecular recognition events that result in the nucleotide sequence of mRNA becoming translated into the amino acid sequence of proteins via the genetic code. The size of an individual gene or an organism's entire genome is often measured in base pairs because DNA is usually double-stranded. Hence, the number of total base pairs is equal to the number of nucleotides in one of the strands (with the exception of non-coding single-stranded regions of telomeres). The haploid human genome (23 chromosomes) is estimated to be about 3.2 billion base pairs long and to contain 20,000–25,000 distinct protein-coding genes. A kilobase (kb) is a unit of measurement in molecular biology equal to 1000 base pairs of DNA or RNA. The total number of DNA base pairs on Earth is estimated at 5.0×1037 with a weight of 50 billion tonnes. In comparison, the total mass of the biosphere has been estimated to be as much as 4 TtC (trillion tons of carbon).

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Wikipedia

DNA

Deoxyribonucleic acid (; DNA) is a polymer composed of two polynucleotide chains that coil around each other to form a double helix. The polymer carries genetic instructions for the development, functioning, growth and reproduction of all known organisms and many viruses. DNA and ribonucleic acid (RNA) are nucleic acids. Alongside proteins, lipids and complex carbohydrates (polysaccharides), nucleic acids are one of the four major types of macromolecules that are essential for all known forms of life. The two DNA strands are known as polynucleotides as they are composed of simpler monomeric units called nucleotides. Each nucleotide is composed of one of four nitrogen-containing nucleobases (cytosine [C], guanine [G], adenine [A] or thymine [T]), a sugar called deoxyribose, and a phosphate group. The nucleotides are joined to one another in a chain by covalent bonds (known as the phosphodiester linkage) between the sugar of one nucleotide and the phosphate of the next, resulting in an alternating sugar-phosphate backbone. The nitrogenous bases of the two separate polynucleotide strands are bound together, according to base pairing rules (A with T and C with G), with hydrogen bonds to make double-stranded DNA. The complementary nitrogenous bases are divided into two groups, the single-ringed pyrimidines and the double-ringed purines. In DNA, the pyrimidines are thymine and cytosine; the purines are adenine and guanine. Both strands of double-stranded DNA store the same biological information. This information is replicated when the two strands separate. The two strands of DNA run in opposite directions to each other and are thus antiparallel. Attached to each sugar is one of four types of nucleobases (or bases). It is the sequence of these four nucleobases along the backbone that encodes genetic information. RNA strands are created using DNA strands as a template in a process called transcription, where DNA bases are exchanged for their corresponding bases except in the case of thymine (T), for which RNA substitutes uracil (U). Under the genetic code, these RNA strands specify the sequence of amino acids within proteins in a process called translation. Within eukaryotic cells, DNA is organized into long structures called chromosomes. Before typical cell division, these chromosomes are duplicated in the process of DNA replication, providing a complete set of chromosomes for each daughter cell. Eukaryotic organisms (animals, plants, fungi and protists) store most of their DNA inside the cell nucleus as nuclear DNA, and some in the mitochondria as mitochondrial DNA or in chloroplasts as chloroplast DNA. In contrast, prokaryotes (bacteria and archaea) store their DNA only in the cytoplasm, in circular chromosomes. Within eukaryotic chromosomes, chromatin proteins, such as histones, compact and organize DNA. These compacting structures guide the interactions between DNA and other proteins, helping control which parts of the DNA are transcribed.

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Wikipedia

TET enzymes

The TET enzymes are a family of ten-eleven translocation (TET) methylcytosine dioxygenases. They are instrumental in DNA demethylation. 5-Methylcytosine (see first Figure) is a methylated form of the DNA base cytosine (C) that often regulates gene transcription and has several other functions in the genome. Demethylation by TET enzymes (see second Figure), can alter the regulation of transcription. The TET enzymes catalyze the hydroxylation of DNA 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), and can further catalyse oxidation of 5hmC to 5-formylcytosine (5fC) and then to 5-carboxycytosine (5caC). 5fC and 5caC can be removed from the DNA base sequence by base excision repair and replaced by cytosine in the base sequence. TET enzymes have central roles in DNA demethylation required during embryogenesis, gametogenesis, memory, learning, addiction and pain perception.

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TOPIC OF THE DAY

Greenwood / Black Wall Street

Before the 1921 destruction of Tulsa’s Greenwood District, Black residents had created a remarkable center of business and community life. The district included stores, professional offices, entertainment venues and homes owned by Black citizens. Understanding Greenwood means learning what was built—not only what was burned.

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TRIVIA QUESTION OF THE DAY

What prosperous Tulsa district became widely known as “Black Wall Street”?

The Greenwood District.