lecture series 9 from dna to protein: genotype to phenotypefrom dna to protein: genotype to...
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Lecture Series 9From DNA to Protein:
Genotype to Phenotype
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From DNA to Protein: Genotype to Phenotype
A. A. Genes and the Synthesis of PolypeptidesGenes and the Synthesis of Polypeptides
B. B. DNA, RNA, and the Flow of Information DNA, RNA, and the Flow of Information akaaka The The Central DogmaCentral Dogma
C. C. Transcription: DNATranscription: DNA--Directed RNA SynthesisDirected RNA Synthesis
D. D. RNA ProcessingRNA Processing
E. E. The Genetic CodeThe Genetic Code
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From DNA to Protein: Genotype to Phenotype
F. F. The Key Players in TranslationThe Key Players in Translation
G. G. Translation: RNATranslation: RNA--Directed Polypeptide SynthesisDirected Polypeptide Synthesis
H. H. Regulation of TranslationRegulation of Translation
I. I. Posttranslational EventsPosttranslational Events
J. J. Mutations: Heritable Changes in GenesMutations: Heritable Changes in Genes
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A. Genes and the Synthesis of Polypeptides
•• Genes are made up of DNA and are Genes are made up of DNA and are expressed in the phenotype as polypeptides.expressed in the phenotype as polypeptides.
•• Beadle and TatumBeadle and Tatum’’s experiments with the s experiments with the bread mold bread mold NeurosporaNeurospora resulted in mutant resulted in mutant strains lacking a specific enzyme in a strains lacking a specific enzyme in a biochemical pathway. These results led to biochemical pathway. These results led to the onethe one--gene, onegene, one--polypeptide hypothesis.polypeptide hypothesis.
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Beadle and Tatum’s evidence for the one gene-one enzyme hypothesis
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A. Genes and the Synthesis of Polypeptides•• Certain hereditary diseases in humans have Certain hereditary diseases in humans have
been found to be caused by a defective been found to be caused by a defective enzyme.enzyme.
•• These observations supported the oneThese observations supported the one--gene, onegene, one--polypeptide hypothesis.polypeptide hypothesis.
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B. DNA, RNA, and the Flow of Information•• RNA differs from DNA in three ways: It is RNA differs from DNA in three ways: It is
singlesingle--stranded, its sugar molecule is ribose stranded, its sugar molecule is ribose rather than rather than deoxyribosedeoxyribose, and its fourth base , and its fourth base is is uraciluracil rather than thymine.rather than thymine.
•• The The central dogmacentral dogma of molecular biology is of molecular biology is DNA DNA →→ RNA RNA →→ protein. Unidirectional when protein. Unidirectional when genes are expressed.genes are expressed.
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B. DNA, RNA, and the Flow of Information •• A gene is expressed in two steps: First, DNA A gene is expressed in two steps: First, DNA
is transcribed to RNA; then RNA is is transcribed to RNA; then RNA is translated into protein.translated into protein.
•• In retroviruses, the rule for transcription is In retroviruses, the rule for transcription is reversed: RNA reversed: RNA →→ DNA. Other RNA viruses DNA. Other RNA viruses exclude DNA altogether, going directly from exclude DNA altogether, going directly from RNA to protein.RNA to protein.
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Overview: the roles of transcription and translation in the flow of geneticinformation (Layer 1)
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Overview: the roles of transcription and translation in the flow of geneticinformation (Layer 2)
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Overview: the roles of transcription and translation in the flow of geneticinformation (Layer 3)
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Overview: the roles of transcription and translation in the flow of geneticinformation (Layer 4)
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Overview: the roles of transcription and translation in the flow of geneticinformation (Layer 5)
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C. Transcription: DNA-Directed RNA Synthesis•• RNA is transcribed from a DNA template RNA is transcribed from a DNA template
after the bases of DNA are exposed by after the bases of DNA are exposed by unwinding of the double helix.unwinding of the double helix.
•• In a given region of DNA, only one of the In a given region of DNA, only one of the two strands can act as a template for two strands can act as a template for transcription.transcription.
•• RNA polymerase catalyzes transcription RNA polymerase catalyzes transcription from the template strand of DNA.from the template strand of DNA.
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C. Transcription: DNA-Directed RNA Synthesis •• Three step process: Initiation, Elongation Three step process: Initiation, Elongation
and Termination.and Termination.•• The initiation of transcription requires that The initiation of transcription requires that
RNA polymerase recognize and bind tightly RNA polymerase recognize and bind tightly to a promoter sequence on DNA.to a promoter sequence on DNA.
•• RNA elongates in a 5RNA elongates in a 5’’--toto--33’’ direction, direction, antiparallelantiparallel to the template DNA. to the template DNA.
•• Special sequences and protein helpers Special sequences and protein helpers terminate transcription.terminate transcription.
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The stages of transcription
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The initiation of transcription at a eukaryotic promoter
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D. RNA Processing
•• After transcription, the preAfter transcription, the pre--mRNA is altered mRNA is altered by the addition of a G cap at the 5by the addition of a G cap at the 5’’ end and end and a poly A tail at the 3a poly A tail at the 3’’ end.end.
•• UTR is UTR is untranslateduntranslated region even though region even though they are transcribed on the mRNA.they are transcribed on the mRNA.
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RNA processing; addition of the 5′ cap and poly(A) tail
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D. RNA Processing
•• The The intronsintrons are removed from the mRNA are removed from the mRNA precursor by the precursor by the spliceosomespliceosome, a complex of , a complex of RNARNA’’s and proteins.s and proteins.
•• These RNAThese RNA’’s are s are snRNAssnRNAs which are ~300 which are ~300 bases long.bases long.
•• Yet another Yet another ribozymeribozyme..
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The roles of snRNPs and spliceosomes in mRNA splicing
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RNA processing: RNA splicing
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Correspondence between exons and protein domains
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E. The Genetic Code
•• The genetic code consists of triplets of The genetic code consists of triplets of nucleotides (nucleotides (codonscodons). Since there are four ). Since there are four bases, there are 64 possible bases, there are 64 possible codonscodons..
•• One mRNA One mRNA codoncodon indicates the starting indicates the starting point of translation and codes for point of translation and codes for methioninemethionine. Three stop . Three stop codonscodons indicate the indicate the end of translation. The other 60 end of translation. The other 60 codonscodonscode only for particular amino acids.code only for particular amino acids.
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E. The Genetic Code
•• Since there are only 20 different amino Since there are only 20 different amino acids, the genetic code is redundant; that is, acids, the genetic code is redundant; that is, there is more than one there is more than one codoncodon for certain for certain amino acids. However, a single amino acids. However, a single codoncodon does does not specify more than one amino acid.not specify more than one amino acid.
•• The genetic code is degenerate but not The genetic code is degenerate but not ambiguous!ambiguous!
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The triplet code
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F. The Key Players in Translation
•• In prokaryotes, translation begins before In prokaryotes, translation begins before the mRNA is completed. the mRNA is completed.
•• In eukaryotes, transcription occurs in the In eukaryotes, transcription occurs in the nucleus and translation occurs in the nucleus and translation occurs in the cytoplasm.cytoplasm.
•• Translation requires three components: Translation requires three components: tRNAtRNA’’ss, activating enzymes, and , activating enzymes, and ribosomesribosomes..
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F. The Key Players in Translation
•• In translation, amino acids are linked in In translation, amino acids are linked in codoncodon--specified order in mRNA. specified order in mRNA.
•• This is achieved by an adapter, transfer RNA This is achieved by an adapter, transfer RNA ((tRNAtRNA), which binds the correct amino acid ), which binds the correct amino acid and has an and has an anticodonanticodon complementary to the complementary to the mRNA mRNA codoncodon..
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The structure of transfer RNA (tRNA)
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The structure of transfer RNA (tRNA)
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F. The Key Players in Translation
•• The The aminoacylaminoacyl--tRNAtRNA synthetasessynthetases, a family , a family of activating enzymes, attach specific amino of activating enzymes, attach specific amino acids to their appropriate acids to their appropriate tRNAtRNA’’ss, forming , forming charged charged tRNAtRNA’’ss..
•• These are the ultimate translators in the These are the ultimate translators in the cell.cell.
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F. The Key Players in Translation
•• The mRNA meets the charged The mRNA meets the charged tRNAtRNA’’ss at a at a ribosome.ribosome.
•• The ribosome is the staging area for protein The ribosome is the staging area for protein synthesis or translation.synthesis or translation.
•• RibosomesRibosomes are roughly 60% RNA and 40% are roughly 60% RNA and 40% proteins.proteins.
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The Ribosome (another ribozyme)
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Translation: the basic concept
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The anatomy of a functioning ribosome
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G. Translation: RNA-Directed Polypeptide Synthesis•• Three step process: Initiation, Elongation Three step process: Initiation, Elongation
and Termination.and Termination.•• An initiation complex consisting of an amino An initiation complex consisting of an amino
acidacid--charged charged tRNAtRNA and a small ribosomal and a small ribosomal subunit bound to mRNA triggers the subunit bound to mRNA triggers the beginning of translation. beginning of translation.
•• Initiation complex includes the use of Initiation complex includes the use of various initiations factors and of 1 GTP.various initiations factors and of 1 GTP.
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The initiation of translation
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G. Translation: RNA-Directed Polypeptide Synthesis •• Polypeptides grow from the N terminus Polypeptides grow from the N terminus
toward the C terminus. The ribosome moves toward the C terminus. The ribosome moves along the mRNA one along the mRNA one codoncodon at a time.at a time.
•• Elongation has three steps: Elongation has three steps: CodonCodonRecognition, Peptide Bond Formation and Recognition, Peptide Bond Formation and Translocation.Translocation.
•• Elongation also requires elongation factors Elongation also requires elongation factors and 3 and 3 GTPsGTPs per amino acid added.per amino acid added.
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The elongation cycle of translation
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G. Translation: RNA-Directed Polypeptide Synthesis •• The presence of a stop The presence of a stop codoncodon in the A site in the A site
of the ribosome causes translation to of the ribosome causes translation to terminate.terminate.
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The termination of translation
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H. Regulation of Translation
•• Some antibiotics work by blocking events in Some antibiotics work by blocking events in translation.translation.
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H. Regulation of Translation
•• In a In a polysomepolysome, more than one ribosome , more than one ribosome moves along the mRNA at one time. moves along the mRNA at one time.
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Coupled transcription and translation in bacteria
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A summary of transcription and translation in a eukaryotic cell
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I. Posttranslational Events
•• Signals contained in the amino acid Signals contained in the amino acid sequences of proteins direct them to cellular sequences of proteins direct them to cellular destinations. destinations.
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I. Posttranslational Events
•• Protein synthesis begins on free Protein synthesis begins on free ribosomesribosomesin the cytoplasm. Those proteins destined in the cytoplasm. Those proteins destined for the nucleus, mitochondria, and plastids for the nucleus, mitochondria, and plastids are completed there and have signals that are completed there and have signals that allow them to bind to and enter destined allow them to bind to and enter destined organelles.organelles.
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I. Posttranslational Events
•• Proteins destined for the ER, Proteins destined for the ER, GolgiGolgiapparatus, apparatus, lysosomeslysosomes, and outside the cell , and outside the cell complete their synthesis on the ER surface. complete their synthesis on the ER surface. They enter the ER by the interaction of a They enter the ER by the interaction of a hydrophobic signal sequence with a channel hydrophobic signal sequence with a channel in the membrane.in the membrane.
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The signal mechanism for targeting proteins to the ER
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I. Posttranslational Events
•• Covalent modifications of proteins after Covalent modifications of proteins after translation include proteolysis, translation include proteolysis, glycosylationglycosylation, , and and phosphorylationphosphorylation. .
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J. Mutations: Heritable Changes in Genes•• Mutations in DNA are often expressed as Mutations in DNA are often expressed as
abnormal proteins. However, the result may abnormal proteins. However, the result may not be easily observable phenotypic not be easily observable phenotypic changes.changes.
•• Raw materials for evolution to operate.Raw materials for evolution to operate.•• Some mutations appear only under certain Some mutations appear only under certain
conditions, such as exposure to a certain conditions, such as exposure to a certain environmental agent or condition.environmental agent or condition.
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J. Mutations: Heritable Changes in Genes •• Point mutations (silent, Point mutations (silent, missensemissense, nonsense, , nonsense,
or frameor frame--shift) result from alterations in shift) result from alterations in single base pairs of DNA.single base pairs of DNA.
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Categories and consequences of point mutations: Base-pair substitution
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Categories and consequences of point mutations: Base-pair insertion or deletion
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The molecular basis of sickle-cell disease: a point mutation
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J. Mutations: Heritable Changes in Genes •• Chromosomal mutations (deletions, Chromosomal mutations (deletions,
duplications, inversions, or translocations) duplications, inversions, or translocations) involve large regions of a chromosome. involve large regions of a chromosome.
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Alterations of chromosome structure
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J. Mutations: Heritable Changes in Genes •• Mutations can be spontaneous or induced. Mutations can be spontaneous or induced. •• Spontaneous mutations occur because of Spontaneous mutations occur because of
instabilities in DNA or chromosomes. instabilities in DNA or chromosomes. •• Induced mutations occur when an outside Induced mutations occur when an outside
agent damages DNA.agent damages DNA.
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