Nucleic Acids - Structure & Function PDF
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Ajman University of Science and Technology
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Summary
This document provides an overview of nucleic acids (DNA and RNA). It details the structure of nucleotides, purines, pyrimidines, and the significant roles of DNA and RNA in biological processes. It covers fundamental aspects of molecular biology.
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Nucleic Acids – Structure & Function The structure of the nucleotide DNA & RNA are long polymers Each nucleotide is a monomer Each nucleotide consists of: 1. Nitrogeneous heterocyclic base Purine Pyrimidine 2. 5 carbon sugar Ribose Deoxyribose 3. Ph...
Nucleic Acids – Structure & Function The structure of the nucleotide DNA & RNA are long polymers Each nucleotide is a monomer Each nucleotide consists of: 1. Nitrogeneous heterocyclic base Purine Pyrimidine 2. 5 carbon sugar Ribose Deoxyribose 3. Phosphoryl group The structure of the nucleotide Both base & sugar have ring structures Sugar numbered with ' prime Base numbered without ' prime Covalent bond between sugar & phosphoryl is phosphoester bond Bond between base & sugar is β-N-glycosidic linkage joining the 1'-C of sugar & N of base The major purine bases Nitrogenous bases are heterocyclic amines Cyclic compounds with at least 1 N atom in the ring Purines are double rings: 6-member ring fused to 5-member ring The major pyrimidine bases Pyrimidines consist of single 6-membered ring The structure of the nucleotide DNA & RNA are polymer of nucleotides (repeating unit of the polymer) Nitrogen base is β-attached to: ribose (RNA) deoxyribose (DNA) The sugar is phosphorylated at carbon 5' The structure of the nucleotide Adenosine triphosphate (ATP) Schematic labels the different portions of the molecule indicating the change as sequential phosphoryl groups are added The structure of DNA & RNA Nucleotides polymerize to from a chain in series of 3' to 5' phosphodiester bonds The 5’-P on one unit esterifies 3’-OH on adjacent unit The terminal 5‘ unit retains the phosphate The backbone of DNA/RNA polymer (in blue) is a sugar-phosphate backbone because it is composed of alternating units of deoxyribose and phosphoryl groups (S-P-S-P-) DNA chain segment 3' carbon is linked to a 5' carbon via a phosphodiester bond The helical structure of DNA DNA consists of two chains of nucleotides coiled around one another in a right-handed double helix S-P-S-P- backbones of two strands spiral outside of the helix like the handrails on a spiral staircase. Nitrogenous bases extend into the center at right angles to the axis of the helix like steps of the spiral staircase The helical structure of DNA The helical structure of DNA The diameter of the helix is 2 nm (20 Å) Complementarity of DNA bases The A-T & G-C base pairs fit the physical dimensions of the helix Purine-Purine = TOO WIDE Purine-Pyrimidine = JUST RIGHT Space between sugar- phosphate backbones Hydrogen bonding of DNA helix Noncovalent H-bonding maintain the double helix between base pairs A forms 2 H bonds with T (A=T) C forms 3 H bonds with G (C≡G) This is called base pairing Diameter of double helix 2 nm Distance dictated by purine-pyrimidine base pairs DNA strands complementarity The two DNA strands are complementary Sequence of bases on one determine the sequence of bases on other strand The two strands of DNA are “antiparallel” – they run in opposite directions This arrangement enables the nitrogenous bases on both strands to come into contact at the center of the molecule Only when the 2 strands are antiparallel can the base pairs form the H-bonds that hold them together Ribbon diagram of double helix DNA 2 strands form right-handed double helix Bases in opposite strands H-bond with AT/GC rule 2 strands are antiparallel 5‘3' & 3’5‘ directions 1 complete 360º turn of the helix, 10 nucleotides 1 complete turn is 3.4 nm and 1 nucleotide is 0.34 nm Prokaryotic chromosomes Prokaryotes - single celled organisms with no membrane bound organelles Chromosomes – DNA pieces with genetic instruction (or genes) In prokaryotes: single chromosome with No true nucleus Chromosome is a circular/loop DNA molecule that is supercoiled, as opposed to the linear eukaryotic chromosomes These “naked” loops of DNA attach to the inner membrane of the prokaryote Nucleoid - region of the cytoplasm where the chromosomal DNA is located. It is not a membrane bound nucleus Prokaryotic chromosomes Eukaryotic chromosomes They are linear, found in the nucleus with variable number & size True nucleus surrounded by a nuclear membrane Nucleosome - a strand of DNA wrapped around a disk of histone proteins DNA appears like beads on a string String of beads coils into a larger 30 nm fiber With additional proteins next coiled in to a 300 nm fiber Eukaryotic chromosomes Eukaryotic chromosomes RNA structure S-P-S-P- backbone for ribonucleotides linked by 3'-5' phosphodiester bonds RNA mainly single-stranded Ribose replaces deoxyribose Uracil replaces thymine Base pairing (A=U) and (C≡G) This H bonding results in portions of the single-strand that become double-stranded (Viruses) DNA replication DNA must be replicated before cell division each daughter cell inherits a copy of each gene Cell missing a critical gene will “die” The process of DNA replication must produce an absolutely accurate copy of the original genetic information Mistakes made in critical genes can result in lethal mutations DNA replication – From structure to function Structure of the DNA molecule suggests a mechanism of accurate replication The replication is catalyzed by DNA polymerase, and enzyme that “read” the nitrogenous bases on one DNA strand adding complementary bases to a newly synthesized strand The product is a new DNA molecule one strand is the original/parent strand, and the other is the newly synthesized strand This strategy is called semiconservative replication The mechanism of DNA replication Semiconservative replication generates 2 new DNA helices Each helix has 2 DNA strands One strand is from the parental DNA The other strand is newly synthesized Bacterial DNA replication Bacterial chromosome is a circular molecule of DNA (~ 3 million nuc.) DNA replication begins at a unique sequence - replication origin Replication moves bidirectionally, 500 nuc/sec Position where new nucleotides are added to the growing daughter strand is the replication fork As DNA synthesis moves bidirectionally, there are two replication forks moving in opposite directions Bacterial DNA replication Details of DNA replication 1st step: separation of the strands 1) Accomplished by helicase, which breaks the hydrogen bonds between base pairs 2) Positive supercoiling results when hydrogen bonds are broken, this is relieved by topoisomerase 3) When supercoiling is relieved, single-strand binding protein binds to the separated strands to keep them apart 4) Primase catalyzes synthesis of a 10-12 base piece of RNA to “prime” the DNA replication Details of DNA replication Involved in early steps Involved in later steps DNA polymerase reaction After the 1st step is completed, DNA polymerase III “reads” the parental strand/template, catalyzing the polymerization of a complementary daughter strand In the polymerization reaction: A pyrophosphate group is released as a phosphoester bond is formed between the 5'-phosphoryl group of the nucleotide being added, and the previous 3'-OH of the nucleotide in the newly synthesized daughter strand Based on the bond formed in the polymerization this is referred to a 5'- 3' DNA polymerase reaction - Details DNA replication – Influencing factors The two DNA strands being replicated are antiparallel to one another DNA polymerase III can only catalyze in the 5’3' direction However, the replication fork moves in one direction with both strands replicated simultaneously Small RNA primers are needed for a starting point of DNA replication RESULT: There are different mechanisms for replication of the two strands The leading strand is replicated continuously Opposite strand (lagging strand), is replicated in segments or discontinuously DNA replication – Leading strand A single RNA primer is produced at the replication origin DNA polymerase III continuously catalyzes the addition of nucleotides in the 5’3' direction DNA replication – Lagging strand Many RNA primers are produced as the replication fork moves along the molecule DNA polymerase III catalyzes the elongation of the new strand in the 5’3' direction As the new strand encounters a previously synthesized new piece synthesis stops at that site Okazaki fragment The process repeats with another primer made at a new location of the replication fork DNA replication – Lagging strand Final steps are: The removal of the RNA primers - DNA polymerase I Filling in the gaps - DNA polymerase I Sealing the fragments into an intact strand of DNA - DNA ligase Replication fork – Detailed view Lagging strand DNA synthesis is more easily visualized here. The DNA polymerase III reads discontinuously & in the opposite direction Information flow in biological systems Central Dogma: “in cells the flow of genetic information contained in DNA is a one-way street that leads from DNA to RNA to protein” Transcription - the process by which a single strand of DNA serves as a template for the synthesis of an RNA molecule (Think of making a COPY) Translation - converts the information from one language of nitrogenous bases to another of amino acids (Think of TRANSLATING into another language) Classes or RNA molecules Messenger RNA (mRNA) A complimentary copy of a gene mRNA has and directs the aa sequence of proteins Ribosomal RNA (rRNA) Structural and functional component of the ribosome Forms ribosomes by reacting with proteins 3 types in prokaryotes & 4 types in eukaryotes Transfer RNA (tRNA) Transfers amino acids to the site of protein synthesis Transfer RNA (tRNA) There is at least one tRNA for each aa to be incorporated into a protein tRNA is single-stranded with typically about 80 nucleotides The overall structure is called a cloverleaf ()البرسيم ورقة Intrachain hydrogen bonding of A=U and C≡G tRNA contains rare bases (minor bases), such as D, T, Y, and ψ The 3′-OH end of the terminal nucleotide has a conserved CCA-3′ sequence to which aa bind Transfer RNA (tRNA) Transcription Transcription is catalyzed by RNA polymerase Produces a copy of only 1 DNA strand Process of transcription has 3 stages: Initiation - RNA polymerase binds to the promoter region of the gene Chain elongation – formation of a 3’5' phosphodiester bond, generating a complementary copy Termination - the final step when the RNA polymerase releases the newly formed RNA molecule Stages of transcription Initiation Elongation Termination Post-transcriptional mRNA processing Prokaryotes release a mature mRNA at the end of termination for transcription Eukaryotic mRNA is a primary transcript that needs post-transcriptional modifications including: A 5' cap structure is added Required for efficient translation of the final mRNA (binding site for ribosomes), and Protect form nucleases-induced mRNA degradation A 3' poly(A) tail (100 to 200 units) is added by poly(A) polymerase It protects the 3' end of the mRNA from enzymatic digestion It makes the mRNA more stable in the cytoplasm and prolongs its life Addition of the 5’-methylated cap to mRNA This cap is recognized by the protein synthesis machinery The 7-methylguanosine links to the 5’ end via 5’/5’-triphosphate link May include additional methylations at 2’-OH groups of the next two nucleotides RNA splicing RNA splicing - removal of portions from primary mRNA that are not protein coding Bacterial prokaryotic chromosomes are continuous - all DNA sequence from the chromosome is found in the mRNA Eukaryotic chromosomes are discontinuous Introns or intervening sequences - extra DNA sequences within the genes that don’t encode for any amino acid RNA splicing RNA splicing must be very precise Removing an incorrect number of nucleotides destroy the code for the protein Signals mark the intron boundaries Spliceosomes - composed of small nuclear ribonucleoproteins (snRNPs) & help: Recognizing intron-exon boundaries Stabilizing the splicing complex RNA splicing schematic diagram 1. Small nuclear RNAs (snRNAs) form a complex called a spliceosome with small nuclear ribonucleoproteins (snRNPs) and other proteins 2. The snRNAs bind to specific nucleotides in the introns of a pre- mRNA. 3. The RNA transcript is cut, releasing The genetic code The properties of the message on DNA that has been translated to mRNA: Degenerate/redundant - more than one three base codon can code for the same aa Specific - each codon specifies a particular amino acid Nonoverlapping & commaless: None of the bases are shared between consecutive codons. The codes are read one after the other on a continuous manner A complete sequence of mRNA, form the initiation to the termination codon, is termed as the open reading frame Universal - all organisms use the same code The genetic code All 64 codons have meaning 61 code for amino acids Three code for the “stop” signal Multiple codes for an amino acid tend to have two bases in common CUU, CUC, CUA, CUG code for leucine Makes the code mutation resistant Codons are written in a 5’ 3' sequence The genetic code Protein synthesis Protein synthesis is called translation Carried out on ribosomes, complexes of rRNA & proteins mRNA is translated 5’3’ producing NC-terminal polypeptide Protein synthesis occurs in multiple places on one mRNA at a time mRNA plus the multiple ribosomes are called a polysome tRNA Binds a specific amino acid aided by aminoacyl tRNA synthetase Recognizes the appropriate codon on the mRNA Schematic of the translation process Ribosomes Ribosomes are complexes of rRNA and proteins Each ribosome is made up of 2 subunits Small ribosomal subunit contains 1 rRNA and 33 proteins Large ribosomal subunit contains 3 rRNA and about 49 proteins Many ribosomes on 1 mRNA comprise a polysome with many copies of the protein made simultaneously Ribosomes – Structure & Function Ribosomes – Structure & Function The role of transfer RNA (tRNA) tRNA - molecules decoding mRNA information into the protein’s primary structure It requires two specific functions: Each tRNA must covalently bind one, only one, specific amino acid Binding site for covalent attachment of amino acid at 3' end Aminoacyl tRNA synthetase covalently links the proper aa to the tRNA aminoacyl tRNA tRNA must recognize the appropriate codon on the mRNA that calls for that aa Aminoacyl tRNA Synthetase It catalyze the attachment of a tRNA molecule to its respective aa There is at least one aminoacyl tRNA synthetase for each aa The attachment of the aa activates/charges the tRNA molecule The attachment of the aa is at its carboxyl terminal Aminoacyl tRNA Synthetase The mechanism of translation Three steps of translation: Initiation - sets the stage for polypeptide synthesis Elongation - causes the sequential addition of aa to the polypeptide chain in a colinear fashion as determined by the sequence of mRNA Termination - brings the polypeptide synthesis to a The mechanism of translation - Initiation The initiation codon is an AUG It determines the reading frame Translation is towards the 5’ end of the mRNA molecule that is being translated Initiation factors (proteins), mRNA, initiator tRNA, and small and large ribosomes come together Ribosome has two sites to bind tRNA P-site binds to the growing peptide The mechanism of translation - Initiation Insert Fig 24.19 The mechanism of translation – Elongation This is a three step process: 1. An aminoacyl tRNA binds to A-site 2. The peptidyl transferase catalyzes the formation of peptide bond 3. Translocation/movement of ribosome down the mRNA chain to next codon Shifts the new peptidyl tRNA from the A-site to the P-site Chain elongation requires hydrolysis of GTP to GDP The mechanism of translation – Elongation Insert Fig 24.19 The mechanism of translation – Termination Upon finding a “stop” codon a release factor binds the empty A- site The bond between the last aa and peptidyl tRNA is hydrolyzed releasing the protein The protein released may not be in its final form Post-translational modifications may occur before a protein is fully functional Cleavage Association with other proteins Bonding to carbohydrate or lipid groups The mechanism of translation – Termination Mutations, UV light, & DNA repair Mutations are mistakes introduced into the DNA sequence of an organism Mutations can be silent no change in the protein sequence Many mutations have a negative effect on the health of the organism Mutagens - chemicals causing a change in the DNA sequence. They are also carcinogens cause cancer Type of mutations – Point mutation Point or substitutional mutation - The most common replacement of one base in the coding strand of DNA with another Different aa Example - In hemoglobin, substitution of one aa result in the fatal sickle cell anemia A point-nonsense mutation results in a premature stop codon, resulting in a truncated, incomplete, and usually nonfunctional protein product Type of mutations – Frameshift mutation Frameshift mutation - One or more bases is/are added or deleted from the normal order of bases in DNA resulting in shifting of all the triplets over by one base Different sequence of aa A deletion mutation occurs when one or more nucleotides is/are lost from a DNA molecule An insertion mutation occurs when one or more nucleotides is/are added to a DNA molecule Type of mutations – Silent mutation Silent mutations occur when the change of a single DNA nucleotide within a protein-coding portion of a gene does not affect the sequence of amino acids that make up the gene's protein UV damage & DNA repair UV causes covalent linkage of adjacent pyrimidine bases Formation of a pyrimidine dimer on a DNA strand Pyrimidine dimer can be used to kill bacteria with UV light UV damage & DNA repair UV damage – Xeroderma Pigmentosum Rare disorder initially described in 1874 It is an UV radiation sensitivity disorder characterized by: Severe skin burning following minimum sun exposure Skin cancer at an early age Recombinant DNA Recombinant DNA is synthetic DNA that contains segments from more than one source joining the two DNA molecules to create a hybrid The technology was made possible by two types of enzymes: restriction endonucleases (bacterial enzymes that cut the backbone of DNA at specific nucleotide sequences) and ligase Three key elements are needed to form recombinant DNA: Recombinant DNA First, bacterial plasmid DNA is cut by restriction endonuclease EcoRI, which cuts in a specific place This gives a double strand of linear plasmid DNA with two ends ready to bond, called sticky ends Recombinant DNA Then, a second sample of human DNA is cut with the same EcoRI This forms human DNA segments with sticky ends that are complimentary to the plasmid DNA Recombinant DNA DNA ligase - combining the two pieces of DNA forms DNA containing the new segment This DNA chain is slightly larger because of its additional segment New DNA is re-inserted into a bacterial cell Recombinant human insulin production Thank you…