Toe-Tapping Transcription and Translation From Gene to Protein... Chapter 17

I. Background Information  DNA contains information for how to “build” an organism  Variations in genes make different phenotypes  Proteins are the links between genotype and phenotype  Proteins are one or more polypeptide  One gene codes for one polypeptide

II. The Big Picture A. Flow of Information Transcription Genes (DNA) mRNA Nucleus Nucleus

II. The Big Picture Translation Translation mRNAProtein/(polypeptide) Ribosome Ribosome

II. The Big Picture B. Difference Between DNA and RNA B. Difference Between DNA and RNA DNARNA SUGARDeoxyriboseRibose BASESA,T,C,GA,U,C,G SIZE Double Strand Single Strand LOCATIONNucleus Leaves nucleus To cytoplasm

II. The Big Picture C. Prokaryotic vs. Eukaryotic  Prokaryotic have no nucleus, so transcription and translation happen simultaneously  Eukaryotic have processes separated, one in nucleus and one in cytoplasm

Genetic Code DNA contains only 4 bases DNA contains only 4 bases There are 20 amino acids There are 20 amino acids To make all 20, you must have combinations of at least 3 nucleotide bases To make all 20, you must have combinations of at least 3 nucleotide bases Triplet Code: DNA contains 3 letter codes = codon that make each amino acid Triplet Code: DNA contains 3 letter codes = codon that make each amino acid

III. Transcription A. The Overview 1. DNA opens up 2. mRNA reads template side (only one side of DNA is used) 3. Don’t forget that Uracil replaces T

III. Transcription B. The Details 1. RNA polymerase opens the two strands of DNA and hooks RNA nucleotides as they base pair along the DNA template 2. Works only in the 5’ to 3’ direction 3. Starts at a promoter sequence. Ends at a terminator sequence Transcription Unit = gene + promoter + terminator 4. As it elongates, strand detaches from DNA template. DNA closes behind it.

III. Transcription Initiation Initiation Promoter – attachment point for RNA polymerase, determines which side of DNA is template Promoter – attachment point for RNA polymerase, determines which side of DNA is template In Eukaryotes – promoter includes a TATA box and transcription factors that help RNA polymerase to bind In Eukaryotes – promoter includes a TATA box and transcription factors that help RNA polymerase to bind

III. Transcription Elongation Elongation RNA polymerase moves along DNA and unwinds it RNA polymerase moves along DNA and unwinds it Brings in correct RNA bases to match DNA template Brings in correct RNA bases to match DNA template RNA peels away from DNA template RNA peels away from DNA template More than one RNA strand can be made at one time More than one RNA strand can be made at one time

III. Transcription Termination Termination Bacteria – when RNA polymerase reaches the termination signal it detaches from DNA and releases the transcript as mRNA Bacteria – when RNA polymerase reaches the termination signal it detaches from DNA and releases the transcript as mRNA Eukaryotes – RNA polymerase reaches polyadenylation sequence AAUAAA and proteins cut off pre-mRNA from the DNA template Eukaryotes – RNA polymerase reaches polyadenylation sequence AAUAAA and proteins cut off pre-mRNA from the DNA template

III. Transcription C. Alteration of mRNA ends (eukaryotes) 1. 5’ end gets a guanine cap (5’ cap) for protection and attach signal for ribosome. 2. 3’ end gets a poly(A) tail ( adenines). This protects, is used for attachment, and aides in transport.

III. Transcription D. RNA splicing 1. Introns = non-coding sections 2. Exons = coding sections Introns get spliced before the mRNA goes to the cytoplasm 3. Spliceosome, made up of small nuclear ribonucleoproteins (snRNPs) and proteins, cut out the introns.

III. Transcription E. Why Introns? Different splicing patterns make different polypeptides Different splicing patterns make different polypeptides Introns may regulate gene activity Introns may regulate gene activity Alternative splicing may make different sections of a protein Alternative splicing may make different sections of a protein

IV. Tricky Translation A. The Basics mRNA (all spliced and modified) goes out of nucleus to cytoplasm to make a protein in the ribosome mRNA (all spliced and modified) goes out of nucleus to cytoplasm to make a protein in the ribosome mRNA is read in groups of three bases called CODONS. mRNA is read in groups of three bases called CODONS. tRNA transfers amino acids from cytoplasm sea to the ribosome tRNA transfers amino acids from cytoplasm sea to the ribosome the ribosome connects the amino acids together the ribosome connects the amino acids together

B. Details on tRNA Each tRNA carries a specific amino acid at one end Each tRNA carries a specific amino acid at one end At the other end is a nucleotide triplet called an anticodon. This base pairs with the mRNA. At the other end is a nucleotide triplet called an anticodon. This base pairs with the mRNA. Made in nucleus, goes to cytoplasm Made in nucleus, goes to cytoplasm Can be used repeatedly Can be used repeatedly Short single strand of nucleotides Short single strand of nucleotides

B. Details on tRNA Folds into 3D shape (we love H-bonds) Folds into 3D shape (we love H-bonds) Wobble - some tRNA’s have anticodons that can recognize two or more codons, these involve switches in the third-position only. Wobble - some tRNA’s have anticodons that can recognize two or more codons, these involve switches in the third-position only. Aminoacyl-tRNA synthetase - Enzyme that joins amino acid to tRNA. 20 varieties, 1 for each amino acid. Aminoacyl-tRNA synthetase - Enzyme that joins amino acid to tRNA. 20 varieties, 1 for each amino acid.

C. Rockin’ Ribosomes 1. Made up of a large and small subunit which are made of proteins and ribosomal RNA 2. Subunits are made in nucleolus 3. Place where mRNA codons and tRNA amino acids come together to make a protein

C. Rockin’ Ribosomes 4. P-site: holds the tRNA carrying the growing polypeptide chain 5. A-site: holds the tRNA carrying the next amino acid to be added to the chain 6. E-site: Discharged tRNA’s leave the ribosome from here.

D. Building a Polypeptide 1. Initiation small ribosomal subunit binds to molecule of mRNA at 5’ end small ribosomal subunit binds to molecule of mRNA at 5’ end An initiator tRNA (methionine-AUG) binds to P- site An initiator tRNA (methionine-AUG) binds to P- site large subunit attaches to make functional ribosome. A-site is ready to get next amino acid large subunit attaches to make functional ribosome. A-site is ready to get next amino acid GTP, form of energy, is expended to put the ribosome together GTP, form of energy, is expended to put the ribosome together

D. Building a Polypeptide 2. Elongation Codon recognition occurs as mRNA in the A- site of ribosome bonds with anticodon of tRNA (with amino acid). This requires GTP. Codon recognition occurs as mRNA in the A- site of ribosome bonds with anticodon of tRNA (with amino acid). This requires GTP. Amino acid in P-site binds to amino acid in A- site with a peptide bond to build the protein. Amino acid in P-site binds to amino acid in A- site with a peptide bond to build the protein. Translocation - ribosome moves tRNA in A-site to P-site. tRNA in P-site is released. Building continues. Translocation - ribosome moves tRNA in A-site to P-site. tRNA in P-site is released. Building continues.

D. Building a Polypeptide 3. Termination Keeps going until stop codon in mRNA reaches A-site of ribosome Keeps going until stop codon in mRNA reaches A-site of ribosome UAA, UAG, UGA are stop codons UAA, UAG, UGA are stop codons Release factor binds to stop codon in A- site. Water added instead of an amino acid Release factor binds to stop codon in A- site. Water added instead of an amino acid

E. Other Fun Stuff 1. Polyribosomes Many ribosomes can work one strand of mRNA to make bulk protein Many ribosomes can work one strand of mRNA to make bulk protein 2. Becoming Functional by Folding Folds into secondary, tertiary, and quaternary structures Folds into secondary, tertiary, and quaternary structures Post translational modifications include attachment of sugars, lipids, phosphates, etc. Post translational modifications include attachment of sugars, lipids, phosphates, etc.

V. Mucky Mutations A. Overview 1. Changes in genetic make-up of a cell 2. Point mutations-Chemical change in just one base pair 3. In gametes, it can go to offspring (sickle- cell anemia)

B. Substitution 1. Replacement of one nucleotide and its partner in the complementary DNA strand with another pair of nucleotides 2. Missense Mutation-altered codon still codes for an amino acid, so it makes sense, just wrong sense 3. Nonsense Mutation-changes amino acid codon to a stop signal – almost always makes nonfunctional protein

C. Insertions or Deletions 1. Additions or losses of nucleotide pairs in a gene 2. Throws off the triplet reading (frame shift) 3. Produces nonfunctional proteins

D. Why/How Mutations? 1. Errors during DNA replication or repair (Spontaneous mutations) 2. Mutagens, like physical and chemical stuff, can interact with DNA and mutate it 3. Mutagens include UV light, X-rays, and chemicals

What is a Gene? A gene is a region of DNA that can be expressed to produce a final functional product that is either a polypeptide or an RNA molecule A gene is a region of DNA that can be expressed to produce a final functional product that is either a polypeptide or an RNA molecule