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One gene, one protein concept
- Transcription
- -Synthesis of mRNA from DNA template (Both nucleic acids use nearly the same code)
- -mRNA transcript of gene's protein instructions (mRNA=messenger RNA)
- Translation
- -Synthesis of polypeptid from mRNA transcript
- -Nucleotides translated into amino acid order
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Triplet code
- mRNA base triplets=codons
- -ex. Codon UGG codes for trytophan
- The code is redundant
- -Reduncancy=1+ triplet codes per amino acid.
- Code is NOT ambiguous
- -Each triplet code is for one amino acid only
- Some triplets do not code for amino acids
- -Stop codons (no amino acid)-termination (end point of translation)
- Start codon (& amino acid methionine)
- initiation point of translation
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Reading Frame
- starting point of reading nucleotides crucial
- ex:
- UUAGCGCCCUAUGCGCCU
- UUAGCGCCCUAUGCGCCU
- There are no spaces in genetic code
- Incorrect starting point results in different polypetptide
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Evolution of Code
- Genetic Code is universal
- Code must have evoled very early
- --Shared by all organisms
- --Very few codon differences
- Universal properties allow gene transfer
- ---insertion of foreign genes in an organism
- ---Biotechnology applications
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Transcription Initiation
- Transcription factors help binding
- -proteins that help start transcription
- Transcription initiation complex
- -Transcription factors and RNA polymerase
- Complex starts transcription at promoter
- -TATA box: specific sequence in promoter
- Two DNA strands unwound
- Transcription is initated.
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Transcription Elongation
- RNA polymerase moves along DNA
- Exposes 10-20 DNA bases for transcription
- RNA polymerase moves on, helix reforms
- Transcription rate ≈ 60 bases / second
- A Gene can be transcribed simultaneously
- –Multiple RNA polymerases on same gene
- –Multiple transcripts produced
- Increases production of a single protein
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DNA termination
- RNA polymerase transcribes terminator
- sequence (AAUAAA)
- Then pre-mRNA is cut free from enzyme
- RNA is further modified after transcription
- –Both ends of the transcript are modified
- Starting point receives a cap molecule
- Cleavage site receives tail sequence
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Modification of pre-mRNA
- Guanosine triphosphate (G-PPP) cap
- -Protects mRNA from degradation
- -Indicates location for ribosome attachment
- Poly A tail (...AAAAAA...) sequence
- -protective
- -aids in export from nucleus
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mRNA splicing
- Not all pre-mRNA transcribed is use
- Sections are cut & pasted = RNA splicing
- Most genes have noncoding nucleotides
- –Exons: coding regions Introns: noncoding
- Primary transcript has both introns & exons
- mRNA that leaves nucleus is reduced
- –Introns removed and exons spliced together
- Done by spliceosomes
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tRNA
- •anticodons bring amino acids
- –tRNA = transfer RNA
- –tRNA transfers amino acids to ribosome
- •From amino acid pool in cytoplasm
- –Ribosome adds amino acids to polypeptide
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Ribosomes
- •Facilitate coupling of tRNA with mRNA
- –Allow matching of anticodons to codons
- •Ribosome consists of two subunits
- –Large subunit, small subunit
- –Subunits formed in nucleoli (nucleus)
- –Exported to cytoplasm via nuclear pores
- •Constructed of rRNA + proteins
- –rRNA = ribosomal RNA
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Initiation
- •Small ribosomal subunit binds to RNAs
- –mRNA and initiator tRNA
- –Attaches to mRNA at 5’ end (leader)
- •Initiator tRNA binds with mRNA codon
- –mRNA initiation codon AUG
- –Initiator tRNA carries methionine
- •Large ribosomal subunit joins complex
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Elongation
- 3 Steps:
- 1)Codon recognition (mRNA in site A binds with incoming tRNA)
- 2) Peptide bond formation (New amino acid joined to polypeptide. Catalyzed by rRNA [acts as ribozhyme] Poly peptide chain moved to newly arrived tRNA)
- 3) Translocation- Ribosome moves newest tRNA from A to P site. Previous tRNA from P site moves to E site. mRNA is moved along with the tRNA
- TERMINATION BRO!
- elongation continues to stop until codon in A site. Release factor protein binds to stop codon. Hydrolysis of polypeptide from tRNA
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Signal peptides
- •Ribosomes can be free or bound
- –Free in cytosol, bound on ER
- –Synthesis of all proteins begins in cytosol
- •Signal peptide is cue for attachment to ER
- •Near leading end of polypeptide
- –Signal recognition particle (SRP)
- •Brings ribosome to receptor on ER
- –Polypeptide released into cisterna of ER
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Base pair substutions, insertions, deletions (mutations)
- –Replacement of one nucleotide pair
- –Can be silent mutations
- •Redundancy of genetic code
- –Could still possibly code for same amino acid
- –Ex. GGC & GGU both code for glycine
- •Or, different amino acid could act similarly
- –Significant if mutation affects protein function
- •Ex. changes active site of enzyme protein
- •Insertions and Deletions
- –Addition or loss of base pairs
- •Change overall length of gene (vs. substitutions)
- –Usually affect gene product much more
- –Creates a frameshift
- •Affects all triplet codons downstream
- •Extensive missense (or nonsense) effect
- •Frameshift usually makes gene nonfunctional
- –Unless frameshift near very end of gene
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