Nucleotides and Nucleic Acids PDF

Nucleotides and nucleic acids Marc A. Ilies, Ph. D. Lehninger - Chapter 8 [email protected]; lab 517 For questions, comments please use the discussion tool in Canvas office 517A (Tu, Fr 3-5)...

Nucleotides and nucleic acids Marc A. Ilies, Ph. D. Lehninger - Chapter 8 [email protected]; lab 517 For questions, comments please use the discussion tool in Canvas office 517A (Tu, Fr 3-5) ©MAIlies2024 1  Nucleotides Roles: - constituents of nucleic acids (e. g. DNA, RNA), the main chemical entities responsible for storing and transferring of genetic information; - mediators of energy storage and release in metabolic processes (e.g. ATP, ADP) - signaling role: essential chemical links for cell’s response to external stimuli (hormones, temperature, pressure) Nucleotides as building blocks of nucleic acids The ability to store and transmit genetic information from one generation to the next one is a fundamental condition of life Requirements: - very stable under a diverse range of environmental conditions - water soluble: water is an intrinsic component of life - presence of recognition (affinity) structural features that can mediate their self-association (self-assembling) 2  Nucleotides: structure - contain three main structural elements: = or Nucleoside Nucleotide 3  Nitrogen-containing bases: purines and pyrimidines - very stable heterocyclic compounds (aromatic character), incorporating atoms/groups that are H-bond donors and acceptors - flat structures, similar to benzene; have π electrons - have tautomers (important for pairing and for chemical reactions mutations): O OH O OH HN N HN N thymine O N O N HO N HO N tautomers H H 4  Nucleosides - Ribose and ribosides: H O NH2 NH2 C N N N N HOCH2 H C OH HOCH2 HOCH2 OH N N N O H O N O H H H H C OH H H H H H OH H H - H2O H H H C OH HO OH HO OH HO OH CH2OH -D-ribofuranose D-ribose -D-ribofuranose 9-(-D-ribofuranosyl)adenine Adenosine - 2-deoxyribose and deoxyribosides: H O NH2 NH2 C N N N N HOCH2 HOCH2 HOCH2 H C H OH N N O H O N N O H H H H C OH H H H H H OH H H - H2O H H H C OH HO H HO H HO H CH2OH 9-(-2-deoxy- -2-deoxy- -2-deoxy- 2-deoxy- D-ribose D-ribofuranosyl)adenine D-ribofuranose D-ribofuranose 5 Deoxyadenosine  Nucleosides: conformation, sugar puckering - relates to various conformations that can be adopted by sugar’s furanosic ring: NH2 N NH2 N N N HOCH2 N HOCH2 O N H N O H N H 1' 3' 1' 2' NH2 H 2' H H HO HH OH N HO HO N HOCH2 N C-2’endo C-3’endo O N H H NH2 H H NH2 N N HO OH N HOCH2 N HOCH2 N N O N O 9-(-D-ribofuranosyl)adenine 1' N H 1' Adenosine 2' 3' 2' H H H H H H HO H HO OH OH C-2’exo C-3’exo 6 Nucleotides 7 8  Other purine and pyrimidine nucleosides - both DNA and RNA contain some minor purine and pyrimidine bases: m-RNA cap 9  Nucleotides: isomers - the phosphate can also be attached on different other positions than 5’: - cyclic esters are also possible, some of them (e.g. 3’,5’- cyclic AMP) have physiologic roles 10  Nucleic acids - DNA and RNA - nucleotides linked through phosphodiester bonds - phosphate group ionized at physiologic pH polyacids - sequence read from 5’ to 3’: Negative charge at pH=7, therefore acidic 11  Nucleic acids: stability - alkaline hydrolysis of RNA is rapid: RNA is a very sensitive molecule, with a limited lifetime: - DNA is resistant because it lacks the 2’-OH’s; DNA fragments of old fossils, mummies can be retrieved intact → anthropology, history - RNA older than DNA; DNA became the preferred molecule for storing genetic information 12 due to its superior stability  Molecular recognition and base pairing in nucleic acids - two antiparallel chains; N-containing bases inside, phosphate backbone outside - base pairs are formed through H bonding (2 for AT, 3 for GC); 13 Discovery of DNA structure 14  Structure of DNA - double helix structure discovered by J.D. Watson, F. Crick, M. Wilkins, R. Franklin - chiral molecule (natural DNA is a right handed double helix) Helix rise per base pair 10.5 basepairs for a full turn diameter 15  DNA forms - Based on the sequence and on the environment we can distinguish several DNA forms: random sequence CG repeats Watson-Crick 16 Form 17  DNA replication 18 https://dnalc.cshl.edu/resources/3d/04-mechanism-of-replication-advanced.html  Unusual DNA structures - double-stranded nucleic acids with - double-stranded nucleic acids with twofold symmetry symmetric sequence within each strand - palindromic DNA or RNA sequences can form alternative structures with intra-strand base pairing: one strand both strands with intra-strand bp with intra-strand bp19  Unusual DNA structures - Several unusual DNA structures involve three or even four DNA strands: Hoogsteen pairing in a form of triplex DNA 20 Triple-helical DNA  Secondary structures of RNA - any self-complementary sequence can produce complex, unique 3D structures: right-handed helical structure of RNA (single stranded !) 21  Transfer –RNA (tRNA) - adapter molecule in protein synthesis - covalently linked to an amino acid at one end, pair with mRNA via anticodon loop - important in translation: tRNA Ribosome 22 mRNA  Unusual base pairs in RNA - the secondary structure of RNA depends heavily on the sequence; note the non-Watson-Crick G=U pairs formed by folding of RNA molecules 23  Messenger–RNA (mRNA) - special structure with several recognition elements: 24  Nucleic acid chemistry (renaturation) DNA hybridization (% of hybrid duplexes reflects a Reversible denaturation of DNA common evolutionary heritage) 25 Heat denaturation of DNA Between G and C there are 3 H-bonds , between A and T there are 2 H-bonds so a lower tm is observed in DNA With many AT’s tm (melting temperature): point at which 50% of helical nature is lost 26 Non-enzymatic reactions of nucleotides - spontaneous deamination of cytosine HNO2 and hydrolysis of purines can occur; - repair mechanisms will normally handle these potentially lethal changes. HNO2 HNO2 HNO2 HNO2 27  Non-enzymatic reactions of nucleotides - Purines more sensitive than pyrimidines to hydrolysis: - 1 in 100,000 lost every day - process accelerated by acidic pH 28  Alkylation of DNA, alkylating agents and DNA damage - methylation of DNA is used in vivo to control gene expression; it is done using S-adenosyl methionine as donor of Me groups: (natural methylator, that can methylate DNA, lipids, proteins, etc) - methylation of DNA in vivo is under strict regulatory control; excessive methylation can cause mutations and cancer: (cannot pair with C) 29  Alkylation of DNA, alkylating agents and DNA damage - we can kill cancer cells by damaging their DNA through methylation /alkylation using chemical alkylating agents: methylators: dialkylators: (have two reactive arms) 30  Alkylating Agents-All are toxic, some used as antineoplastics Cl H3C N H 3C N Mechlorethamine Nitrogen Cl Cl Mustard O HN N H2N N N H3C N DNA chain Cl O O CH3 N N HN N NH Cl N N NH2 N H2N N DNA polymerase DNA Chain DNA Chain cannot repair the alkylated strands Cross Linked DNA chains by alkylation with Nitrogen Mustard CH3 ' O G C O N CH3 → N N HN NH N N N N NH2 H2N N C G 3' 31 DNA Chain DNA Chain  Interacting with DNA in cancer & oncology Terminology: - affected tissues: - malign(ant), malignancies, neoplasms, tumors - benign – non cancerous - drugs: “oncologic drugs, oncolytics, antineoplastics, cytotoxic agents, chemotherapeutic agents (Chemo), anti-cancer agents; - several classes of chemotherapeutic agents (detailed in PY2): 32  Inhibitors of DNA transcription - intercalate between two successive GC base pairs, bending the DNA and blocking the transcription through blocking of RNA polymerase: Stabilizing structures (make H bonds with backbone) - anticancer agent doxorubicin acts through this mechanism 33  Damage of DNA by UV radiation - energy-rich radiation (UV, X-rays, etc) are causing chemical changes in DNA via photochemical reactions - radiations are believed to be responsible for 10% of the total DNA damage caused by environmental agents Xeroderma pigmentosum: lack of UV-specific endonuclease which repairs thymine dimers 34  Other functions of nucleotides 1. Energy storage ATP: Energy currency of life (similar energy-rich molecules) 35  Other functions of nucleotides 2. Redox coenzymes - associated with dehydrogenases 36  Other functions of nucleotides 2. Redox coenzymes - associated with dehydrogenases 37  Other functions of nucleotides 3. Regulatory Nucleotides 38  Goals and Objectives Upon completion of this lecture at minimum you should be able to answer the following: ►What is the structure of nucleotides and what roles can the play in vivo? ►What is nucleoside? What is a nucleotide? What N-containing bases can be found in the structure of nucleoside and nucleotides and which are their main characteristics? ►Which nucleotides are found in the DNA and which ones can be found in RNA? ►Which nucleosides can be found in smaller percentage in the structure of DNA and RNA, and what is their role? ►What phosphate isomers can be distinguished for various nucleotides and what is their role in vivo? ►What is the structure of DNA and RNA? What are the main differences between the two main types of nucleic acids? What is fundamentally different in their chemical behavior? ►What are the main differences between A, B, and Z DNA forms? ►What types of unusual DNA structures can we encounter and how are they generated? ►What types of secondary structures can be observed for RNA? ►What are the main structural features of mRNA? ►What is DNA reversible denaturation, what parameters can be defined for the process and what kind of information about DNA sequence can be extracted? ►Which are the main non-enzymatic reactions of nucleotides, what agents are involved, and which are the physiological consequences of these reactions? ►How can DNA be damaged by UV radiation and what disease is associated with this process? ►Besides being constituents of nucleic acids, what other functions are observed for nucleotides, where, and what is the structure of those compounds ? 39  Drugs and diseases ► Diseases: Cancer, Xeroderma pigmentosum ► Drugs: Anticancer drugs (chemotherapeutic agents): alkylators, nitrogen mustards (mechlorethamine), intercalators (doxorubicin), pyrimidine analogs (5-fluorouracil), purine analogs (6-mercaptopurine) 40