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Chapter 9 Proteins and Their Synthesis. Green Fluorescent Protein drawn in cartoon style with fluorophore highlighted as ball-and-stick; one wholly-reproduced protein, and cutaway version to show the fluorophore. Review Central Dogma. 5 ’ ATG GAC CAG TCG GTT TAA GCT 3 ’
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Chapter 9 Proteins and Their Synthesis Green Fluorescent Protein drawn in cartoon style with fluorophore highlighted as ball-and-stick; one wholly-reproduced protein, and cutaway version to show the fluorophore.
ReviewCentral Dogma 5’ ATG GAC CAG TCG GTT TAA GCT 3’ 3’ TAC CTG GTC AGC CAA ATT CGT 5’ DNA RNA Protein transcription 5’AUG GAC CAG UCG GUU UAA GCU 3’ translation aa - aa - aa - aa - aa - aa - aa
Protein Structure via condensation
Protein Structure Primary Structure
Protein folding is dependent on the amino acid R groups R H2N C COOH H General Structure There are 20 amino acids. Their properties are determined by the R group.
There are 20 amino acids. • Nonpolar or hydrophobic (9) • Polar (hydrophillic), but uncharged (6) • Polar (hydrophillic), but charged (5)
Nonpolar (Hydrophobic) ring
Protein Structure Primary Structure
Protein Structure Two major types of Secondary Structure α Helix β Sheet
How do we get from DNA to Primary protein structure ? 5’ ATG GAC CAG TCG GTT TAA GCT 3’ 3’ TAC CTG GTC AGC CAA ATT CGT 5’ DNA RNA Protein transcription 5’AUG GAC CAG UCG GUU UAA GCU 3’ translation aa - aa - aa - aa - aa - aa - aa
DNA (mRNA) is read in Triplets -Codon – Group of 3 DNA bases codes for a specific amino acid Ex. ATG = methionine -This means the code is degenerate – more than one codon can specify one amino acid
Key To The Genetic Code Groups of 3 mRNA bases (codons) code for specific amino acids 5’ CCAACCGGG 3’ CCA-ACC-GGG Pro-Thr-Gly
The Genetic Code – Stop Codons UGA UAA UAG
Proteins and Genes are Colinear Mutations in DNA show specific corresponding changes in the protein Genes are converted to proteins in a linear fashion
Key To The Genetic Code CCG UGG AGA GAC UAA Pro – Trp – Arg –Asp - Stop CCG UCG AGA GAC UAA Pro – Ser – Arg –Asp - Stop CCG UGG CGA GAC UAA Pro – Trp – Arg –Asp - Stop CCG UGG AGA GAC UAA Pro – Stop CCG UGG AGA CGA CUA Pro – Trp – Arg –Arg - Leu
The Genetic Code - Mutations • 4 Types of Mutations • Silent mutations • Missense mutations • Nonsense mutations • Frameshift mutations
The Genetic Code mRNA has 3 potential “reading frames” 5’ CUUACAGUUUAUUGAUACGGAGAAGG 3’ 3’ GAAUGUCAAAUAACUAUGCCUCUUCC 5’ 5’CUU ACA GUU UAU UGA UAC GGA GAA GG 3’ 3’GAA UGU CAA AUA ACU AUG CCU CUU CC 5’ 5’ C UUA CAG UUU AUU GAU ACG GAG AAG G 3’ 3’ G AAU GUC AAA UAA CUA UGC CUC UUC C 5’ 5’ CU UAC AGU UUA UUG AUA CGG AGA AGG 3’ 3’ GA AUG UCA AAU AAC UAU GCC UCU UCC 5’ Stop UAA UGA UAG
The Genetic Code mRNA has 3 potential reading frames 5’ CUUACAGUUUAUUGAUACGGAGAAGG 3’ 3’ GAAUGUCAAAUAACUAUGCCUCUUCC 5’ 5’ CUU ACA GUU UAU UGA UAC GGA GAA GG 3’ 3’ GAA UGU CAA AUA ACU AUG CCU CUU CC 5’ 5’ C UUA CAG UUU AUU GAU ACG GAG AAG G 3’ 3’ G AAU GUC AAA UAA CUA UGC CUC UUC C 5’ 5’ CU UAC AGU UUA UUG AUA CGG AGA AGG 3’ 3’ GA AUG UCA AAU AAC UAU GCC UCU UCC 5’ Stop UAA UGA UAG
Review - RNA mRNA- messenger RNA tRNA- transfer RNA rRNA- Ribosomal RNA
tRNA-The adapter • -tRNA functions as the adapter between amino acids and the RNA template • -tRNAs are structurally similar except in two regions • Amino acid attachment site • Anticodon
tRNA-The anticodon • The tRNA anticodon • 3 base sequence • Complementary to the codon • Base pairing between the mRNA and the tRNA • Oriented and written in the 3’ to 5’ direction tRNA mRNA 3’ CUG 5’ 5’ GAC 3’ Aspartic Acid
Aminoacyl-tRNA synthetase The enzyme responsible for joining an amino acid to its corresponding tRNA 20 tRNA synthetases – 1 for each amino acid
Wobble Allows one tRNA to recognize multiple codons Occurs in the 3rd nucleotide of a codon
Wobble – A new set pairing of rules I = Inosine: A rare base found in tRNA
Wobble – A new set pairing of rules Isoaccepting tRNAs: tRNAs that accept the same amino acid but are transcribed from different genes
Wobble Problem What anticodon would you predict for a tRNA species carrying isoleucine?
Ribosomes – General characteristics • Come together with tRNA and mRNA to create protein • Ribosome consist of one small and one large subunit • In prokaryotes, 30S and 50S subunits form a 70S particle • In Eukaryotes, 40S and 60S subunits form an 80S particle • Each subunit is composed of 1 to 3 types of rRNA and up to 49 proteins
Ribosomes – General characteristics • rRNA folds up by intramolecular base pairing
Translation Synthesizing Protein
Translation Initiation - Prokaryotes Translation begins at an AUG codon – Methionine Requires a special “initiator” tRNA charged with Met – tRNAMeti This involves the addition of a formyl group to methionine while it is attached to the initiator
Shine-Dalgarno Sequence mRNA only associates with unbound 30S subunit
Translation Initiation – ProkaryotesInitiation Factors 3 initiation factor proteins are required for the start of translation in prokaryotes IF1 – Binds to 30S subunit as part of the complete initiation complex. Could be involved in stability IF2 – Binds to charged initiator tRNA and insures that other tRNAS do not enter initiation complex IF3 – Keeps the 30S subunit disassociated from the 50S subunit and allows binding of mRNA
Figure 2-12-1 Figure 9-15-1
Figure 2-12-1 Figure 9-15-2
Figure 2-12-1 Figure 9-15-3
Translation Initiation – Eukaryotes • mRNA is produced in the nucleus and transported to the cytoplasm • 5’ end of the mRNA is “capped” to prevent degradation • Eukaryotic Initiation Factors (eIF4A, eIF4B, and eIF4G) associate with the 5’ cap, the 40S subunit, and initiator tRNA • Complex moves 5’ to 3’ unwinding the mRNA until an initiation site (AUG) is discovered • Initiation factors are released and 60S subunit binds
Figure 2-12-1 Figure 9-16-1 • mRNA is produced in the nucleus and transported to the cytoplasm • mRNA is covered with proteins and often folds on itself • 5’ end of the mRNA is “capped” to prevent degradation
Figure 2-12-1 Figure 9-16-2 4.Eukaryotic Initiation Factors (eIF4A, eIF4B, and eIF4G) associate with the 5’ cap, the 40S subunit, and initiator tRNA
Figure 9-16-3 5. Complex moves 5’ to 3’ unwinding the mRNA until an initiation site (AUG) is discovered
Figure 9-16-4 6. Initiation factors are released and 60S subunit binds
Elongation • Requires two protein Elongation Factors: • EF-Tu and EF-G • Amino acids are added to the growing peptide chain at the rate of 2-15 amino acids per second