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From Gene to Protein Lecture 14 Fall 2008
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Function of DNA Genotype Phenotype Sequence of nucleotide bases in DNA
1 Genotype Sequence of nucleotide bases in DNA All the alleles of every gene present in a given individual Gene : a discrete unit of hereditary information consisting of a specific nucleotide sequence in DNA Phenotype Any observable traits in an individual Physical, physiological & behavioral
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Function of DNA How does the genotype produce the phenotype?
2 How does the genotype produce the phenotype? Gene expression Creates proteins from DNA “one gene – one enzyme” “one gene – one protein” The function of a gene is to dictate the production of a polypeptide Fig.17.3
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Function of DNA Two processes: Transcription Translation
3 Two processes: Transcription The transfer of genetic information from DNA into RNA Synthesis of RNA Nucleus Translation The transfer of information from RNA into proteins (via the ribosomes) Synthesis of polypeptide Cytoplasm Fig. 17.3
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Protein Structure Primary structure Polypeptide chain
4 Primary structure Polypeptide chain Unique sequence of amino acids Amino acids 20 types Basic structure of each amino acid is the same Unique side group = R group See Fig. 5.17
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Overview of Transcription & Translation
5 Transcription “rewrites” DNA into RNA Still in nucleic acid “language” Translation Converts nucleic acid language into polypeptide (amino acid) language Fig. 17.4
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Overview of Transcription & Translation
6 Triplet code of DNA Non-overlapping sequence of 3 bases that code for amino acid 64 possible combinations (43) RNA complementary to DNA Fig. 17.4
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The Dictionary of the Genetic Code
7 Codons mRNA triplets 64 possible combinations (43) Only 20 amino acids Redundancy Start code Stop code Fig. 17.5
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Transcription Transcription
8 Transcription The transfer of genetic information from DNA into RNA 3 main steps Initiation of Transcription RNA Elongation Termination of Transcription
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Transcription Initiation DNA strand separates at promoter
9 Initiation DNA strand separates at promoter Specific area of DNA that designates the start of a gene Contains start point and several dozen nucleotide pairs Determines template strand RNA polymerase binds at promoter Unwinds DNA strand & joins nucleotides RNA synthesis begins Nucleotides that will form RNA line up with DNA nucleotides U replaces T in the RNA strand Bases of RNA & DNA joined by hydrogen bonds Doesn’t need a primer 5’to 3’ Fig. 17.7
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Transcription Transcription Factors
10 Transcription Factors Mediate binding of RNA polymerase to promoter & initiation (eukaryotes) One TF binds to TATA box Transcription initiation complex Complex of RNA polymerase and transcription factors Fig. 17.8
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Transcription RNA Elongation RNA synthesis continues
11 RNA Elongation RNA synthesis continues ~10-20 DNA bases exposed at a time ~ 40 nucletides/second (eukaryotes) “peels off” of DNA DNA strands come back together after RNA removed Fig. 17.7
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Transcription Termination of Transcription Bacteria Eukaryotes
12 Termination of Transcription Bacteria RNA polymerase reaches terminator sequence Signals the end of the gene RNA polymerase detaches from DNA & RNA Eukaryotes Polyadenylation signal sequence DNA segment (UAAUAAA) RNA cut ~10-35 nucleotides downstream of signal RNA polymerase continues transcribing until stopped by enzyme Fig. 17.7
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RNA Processing Primary transcript (pre-mRNA) altered
13 Primary transcript (pre-mRNA) altered Addition of 5” cap and poly-A tail 5’ cap Modified guanine nucleotide Occurs soon after RNA synthesis begins Poly-A tail adenine nucleotides Function Facilitate export from nucleus Protection from degradation by hydrolytic enzymes Facilitate attachment of ribosome to 5’ end Untranslated regions (UTR) Fig. 17.9
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RNA Processing RNA splicing Removal of introns (intervening sequences)
14 RNA splicing Removal of introns (intervening sequences) Intron: non-coding regions of nucleotides Joining of exons (expressed sequences) Exons : coding regions of nucleotides Exceptions – UTRs (untranslated regions) Average polypeptide = 400 (1200 base pairs) Average transcription unit = 27,000 base pairs Results in mRNA (messenger RNA) which then leaves the nucleus Fig
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RNA Processing Spliceosome
15 Spliceosome Complex of small nuclear ribonucleoproteins (snRNPs) and other proteins Binds w/pre-RNA Intron carries region recognized by snRNPs (snRNA) Cuts out introns Binds exons together Fig
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RNA Processing Ribozymes Three characteristics
16 Ribozymes RNA molecules that function as enzymes RNA splicing In some organisms Three characteristics Single strand allows RNA to base pair with itself to form particular 3-D structure Some functional groups on bases can act in catalysis Ability to hydrogen bond with other nucleic acids adds specificity of catalytic activity
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Translation Translation
17 Translation Translation Conversion of nucleic acid language into polypeptide (amino acid) language Synthesis of a polypeptide 3 main steps Initiation Elongation Termination
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Translation Transfer RNA (tRNA)
18 Transfer RNA (tRNA) Converts the codon of mRNA into an amino acid Anticodon at one end Complementary pairing with codon on mRNA Amino acid attachment site at other end Fig
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Translation Attachment of amino acid to tRNA More codons than tRNAs
19 Attachment of amino acid to tRNA ATP Aminoacyl-tRNA synthetases 20 types More codons than tRNAs ~45 different tRNAs “wobble” Some tRNAs able to bind to more than one codon 3rd base has flexibility Fig
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Translation Ribosomes are site of translation
20 Ribosomes are site of translation Made of proteins & ribosomal RNA (rRNA) Small & large subunits Made in nucleolus Exported to cytoplasm Aminoacyl-tRNA binding site Peptidyl-tRNA binding site Exit site Tetracycline and streptomycin target bacteria ribosomes Fig
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Translation Initiation Translation initiation complex formed
21 Initiation Translation initiation complex formed mRNA binds to small subunit Initiator tRNA (UAC anticodon) binds to mRNA at start codon (AUG) Large ribosomal subunit binds to small subunit tRNA positioned in P site Requires initiation factors & GTP Tetracycline and streptomycin target bacteria ribosomes Fig
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Translation Elongation Requires elongation factors & GTP
22 Elongation Requires elongation factors & GTP Codon recognition Peptide bond formation Catalzed by rRNA part of large ribosomal subunit Translocation Fig
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Translation Termination When stop codon on mRNA reaches the A site
23 Termination When stop codon on mRNA reaches the A site UAG, UAA, UGA Release factors bind to stop codon at A site Adds H2O molecule to break bond between polypeptide and tRNA in P site Polypeptide chain free Fig
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Translation Polyribosomes
24 Polyribosomes Multiple ribosomes translating same strand of mRNA simultaneously Increases speed of translation Fig
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From Genotype to Phenotype
25 Fig
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Protein Completion 26 Poypeptide chain folds spontaneously as it is being synthesized Chaparonins Post-translational modifications Additions of sugars, lipids, phosphate groups Removal of amino acids from end of polypeptide chain Quaternary structure formed
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Targeting Proteins Free & bound ribosomes
27 Free & bound ribosomes Polypeptide synthesis always begins in cytosol Signal peptide Sequence of ~ 20 amino acids at leading end of polypeptide Targets protein for ER Signal-recognition particle (SRP) Brings ribosome to translocation complex Protein complex in ER with recognition proteins – forms pore Fig
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Mutations Mutation Change in the nucleotide sequence of DNA
28 Mutation Change in the nucleotide sequence of DNA May give rise to an altered protein Point mutations Chemical changes in a single base pair of a gene Fig
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Types of Point Mutations
29 Base-pair substitutions Replacement of one nucleotide and its partner with another nucleotide pair Silent mutation No effect on polypeptide structure Redundancy in codons
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Types of Point Mutations
30 Missense mutation Changes one amino acid to another May not affect function of protein May significantly alter protein function Nonsense mutations Changes amino acid codon to a stop codon Terminates translation early Nonfunctional protein
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Types of Point Mutations
31 Insertions or deletions Frameshift mutation Alters reading frame Fig
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Mutations Can be spontaneous Can be caused by mutagen
32 Can be spontaneous Errors during DNA replication or recombination Rare: In DNA replication, only 1 in a billion bases incorrectly paired Cell has some repair mechanisms Can be caused by mutagen Physical or chemical agent that causes change E.g., x-rays, UV light, agent orange, tobacco Spontaneous mutations are rare, but rate of mutation increased by mutagens
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Mutations and Evolution
33 Mutations and Evolution If mutation occurs in somatic cells, the mutation is not passed on to offspring If mutation occurs in cells that produce gametes, mutation can be passed on to offspring Mutations are one way of introducing new alleles Increases genetic diversity in populations
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