8/13/2016 Central Dogma of Biology Chapter 17 Flow of genetic information: PROTEIN SYNTHESIS: FROM GENE TO PROTEIN RNA has many functions in the cell. RNA (ribonucleic acid) DIFFERENCES 1. pre-mRNA: precursor to mRNA, newly transcribed and not edited DNA RNA 2. mRNA: carries the code from DNA that specifies amino acids ribose sugar deoxyribose sugar 3. tRNA: carries a specific amino acid (anticodon)to ribosome double strand single strand 4 . rRNA: makes up 60% of the ribosome; site of protein synthesis bases A,T,C,G bases A, U ,C,G 5. snRNA: small nuclear RNA; part of a spliceosome (in RNA splicing) found in: found in: nucleus, 6. srpRNA: a signal recognition particle that binds to signal peptides nucleus, mitochondria, cytosol, chloroplasts ribososomes 7. RNAi: interference RNA; a regulatory molecule many functions 8. ribozyme: RNA molecule that functions as an enzyme PROTEIN SYNTHESIS/GENE EXPRESSION 3 types RNA directly involved in making proteins 1. messenger RNA (mRNA) Formation of proteins using information coded on DNA and single uncoiled long strand carried out by RNA. - transmits DNA info during protein synthesis one gene = one RNA molecule - serves as template to assemble amino acids Gene: region of DNA whose final product is either a polypeptide or RNA molecule 2. transfer RNA (t RNA) - carries amino acids to ribosome A gene is expressed when protein synthesis is occurring. 3. ribosomal RNA (r RNA) makes up large part (2/3) of ribosome - globular 1
8/13/2016 The Genetic Code Code word/Codon (triplet): How is information necessary for creating proteins encoded in the specific group of 3 successive bases on DNA and mRNA RNA? - codes for a specific amino acid to be placed on The genetic code from DNA is transcribed onto m RNA by Codons . the protein chain For each gene, one DNA strand - 20 biological amino acids, but more than 20 codons is the template strand Like “genetic words” DNA code words: ACT, GCA, TTA mRNA (5 ’ 3 ’ ) complementary RNA codons: U GA, CG U , AA U to template mRNA triplets ( codons ) code for amino acids in polypeptide chain How many combinations of code words can we make from 4 bases? 64 different codon combinations ( 4 3 = 64) ** each code word always codes for same amino acid** Redundancy : 1+ codons code for each of 20 AAs Reading frame : groups of 3 must be read in correct groupings Ala: Alanine Cys: Cysteine Asp: Aspartic acid Glu: Glutamic acid This code is universal: all life forms use the same code Phe: Phenylalanine Gly: Glycine His: Histidine Ile: Isoleucine Lys: Lysine Leu: Leucine Met: Methionine Asn: Asparagine Pro: Proline Gln: Glutamine Arg: Arginine Ser: Serine Thr: Threonine Val: Valine Trp: Tryptophane Tyr: Tyrosisne It is possible for non-Watson-Crick base pairing to occur at the third codon position. How do these code words affect protein synthesis? This has phenomenon been termed the wobble hypothesis. Order of code words codes for Order of amino acids codes for Specific type of protein 2
8/13/2016 BUILDING OF PROTEINS 2 Stages of Protein Synthesis - DNA unzips - original strand of DNA acts as template for m RNA (only one strand of DNA molecule needed for transcription) euchromatin : uncoiled areas of DNA, active site of transcription antisense /non-template strand: non coding strand of DNA sense/template strand: coding strand of DNA Stages of Protein Synthesis Transcription 1. Initiation (prokaryotes) I. Transcription (nucleus) nontemplate strand RNA polymerase binds template strand directly to promoter in Process where mRNA is produced from DNA DNA Transcription unit : stretch of DNA that - promoter : region of DNA that initiates transcription of a particular gene. codes for a polypeptide or RNA - located near the transcription start sites of genes on sense strand - located upstream on the DNA (towards the 5’ region of the antisense strand) **can be about 100 – 1000 base pairs long** Steps of transcription 2. Elongation 1. Initiation (eukaryotes) A. RNA polymerase reads DNA template strand A. The promoter site on the B. Complementary nucleotides are added to the 3' DNA contains a sequence end of RNA using information in DNA called a TATA box as instructions - TATAAAA - recognized by RNA polymerase ll **Polymerases always work from the 3' - can be up to 25 bases to the 5' end of the coding strand of DNA (template); thus the upstream from point of antiparallel structure it is forming is transcription going from the 5' to 3’ direction. B. Transcription factor proteins and RNA polymerase ll bind C. As RNA polymerase moves, it untwists to promoter section of DNA DNA, then rewinds it after mRNA is made and unwinds the part of the DNA to be transcribed. D. Once RNA nucleotides are attached to DNA chain, codons are in proper order (this is the structural gene: codes for a single protein) 3
8/13/2016 RNA Processing Occurs After Transcription 3. Termination 1. Alteration of pre-mRNA ends A. RNA polymerase transcribes a terminator sequence in DNA - 5’ cap (modified guanine) and 3 ’ poly-A tail (50-520 A ’ s) are added B. mRNA and polymerase detach. * prokaryotes- mRNA ready - functions: to use • help export mature mRNA from nucleus • protect mRNA from enzyme degradation * eukaryotes- pre-mRNA, • help ribosomes attach to mRNA will undergo further modifications transcription animation RNA Processing Occurs After Transcription RNA Splicing, cont. 2. RNA Splicing snRNPs s mall n uclear r ibo n ucleo p roteins - Pre-mRNA has introns (noncoding sequences) and exons (codes for amino acids) snRNP = snRNA + protein - Splicing - introns cut out, exons joined together Pronounced “ snurps ” Recognize splice sites snRNPs join with other proteins to form a spliceosome Spliceosomes catalyze the process of removing introns and joining exons - Once RNA is spliced it can move out of the nucleus to be RNA splicing animation translated Ribozyme = RNA acts as enzyme Prokaryotic vs Eukaryotic Gene Expression Importance of Introns Prokaryotes Eukaryotes • Functions not known for most introns • Transcription and translation both • Transcription in nucleus; in cytoplasm translation in cytoplasm • Some regulate gene expression • RNA poly binds directly to • DNA in nucleus, RNA travels promoter in/out nucleus • No introns • Alternative RNA Splicing : produce • RNA poly binds to TATA box & • No RNA splicing different combinations of exons transcription factors • RNA contains introns • Depends which segments are • RNA splicing treated as exons during splicing • One gene can make more than one polypeptide • 20,000 genes 100,000 polypeptides Changes in gene expression may confer an evolutionary advantage 4
8/13/2016 Translation (in cytoplasm at ribosome) Transfer RNA (t RNA) - function: transfers amino - process whereby protein is synthesized Aminoacyl-tRNA-synthetase : enzyme acids to ribosome that binds tRNA to specific amino from mRNA acid - 20 types – one for each amino acid (specific) - newly synthesized mRNA moves from nucleus to ribosome in cytoplasm - structure (cloverleaf) - gene has 3x more nucleotides than the protein it - found in cytosol makes Ex: 100 a.a. = 300 nucleotides - components of translation mRNA- message • tRNA- interpreter • ribosome- site of translation • Steps of translation Ribosomes - made in nucleolus 1. Initiation A. 5’ end of m RNA binds to small subunit - 2 subunits make up B. initiator tRNA carrying Met attaches to P site ribosome C. large subunit attaches (ribosome ready for protein synthesis) - about 2/3 is r-RNA and - sites: locations on ribosome where tRNA anticodons 1/3 is protein attach P (peptidyl) site- holds aa chain - smaller subunit has A (aminoacyl) site- holds aa to be binding site for mRNA added E (exit) site- exit for tRNA - normally apart in cytoplasm, come * start codon ( AUG ) will be at the site together during on mRNA where this occurs protein synthesis ** anticodon on first tRNA will always be UAC, amino acid 1 will always be methionine 2. Elongation 3. Termination A. t-RNA with a specific anticodon binds a A. stop codon is reached (UAA, UGA, or UAG). specific amino acid. This happens for several t-RNAs and proper corresponding amino acids in the cytoplasm. B. release factor binds to stop codon and polypeptide is released ATP : energy source used to bind the amino acid to the t-RNA. Aminoacyl-tRNA synthase : enzyme C. subunits dissociate (can be used over again) that does the binding. B. First tRNA binds to P site, second tRNA D. protein is released into cell binds to A site (anticodons are complementary to mRNA codons) E. mRNA is broken down by cell (not be used again – only once) C. Peptidyl transferase reaction occurs: #1 a.a. joins to #2 a.a. F. tRNA is released into cell (used over again) D. Ribosome moves down mRNA and first tRNA is released to be used over again - translocation : movement of ribosome down mRNA E. Amino acids continue to be added to protein chain thru same mechanism - peptidyl synthase : enzyme that joins a.a. together 5
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