From Gene to Protein. How Genes Work. AP Biology
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1 From Gene to Protein How Genes Work
2 What do genes code for? How does DNA code for cells & bodies? how are cells and bodies made from the instructions in DNA DNA proteins cells bodies
3 The Central Dogma Flow of genetic information in a cell How do we move information from DNA to proteins? DNA RNA protein trait replication DNA gets all the glory, but proteins do all the work!
4 Transcription from DNA nucleic acid language to RNA nucleic acid language
5 RNA ribose sugar N-bases uracil instead of thymine U : A C : G single stranded lots of RNAs mrna, trna, rrna, sirna DNA transcription RNA
6 Transcription = Making mrna transcribed DNA strand = template strand untranscribed DNA strand = coding strand same sequence as RNA synthesis of complementary RNA strand transcription bubble Enzyme is RNA polymerase coding strand 5 DNA C G 3 A G A T T C T A rewinding G C T A G G C C C G A A T T U A C C G G G C T U A A 3 T T A C G A C T A G T A T unwinding 3 5 build RNA 5 3 mrna 5 RNA polymerase template strand
7 RNA polymerases 3 RNA polymerase enzymes RNA polymerase 1 only transcribes rrna genes makes ribosomes RNA polymerase 2 transcribes genes into mrna RNA polymerase 3 only transcribes trna genes each has a specific promoter sequence it recognizes
8 Which gene is read? Promoter region binding site before beginning of gene TATA box binding site binding site for RNA polymerase & transcription factors Enhancer region binding site far upstream of gene turns transcription on HIGH
9 TRANSCRIPTION = DNA RNA Occurs in - the NUCLEUS in EUKARYOTES; - in cytoplasm in PROKARYOTES
10 1. INITIATION RNA POLYMERASE binds to DNA at region called PROMOTER LIKE DNA POLYMERASE: can only attach nucleotides in 5 3 direction; UNLIKE DNA POLYMERASE: can start a chain from scratch; no primer needed In eukaryotes: TRANSCRIPTION FACTORS & TATA BOXES help position/bind to correct spot RNA POLYMERASE separates the DNA strands to begin transcription
11 2. ELONGATION RNA chain grows in the 5' 3' direction nucleotides base pair with template strand; nucleotides added to the 3' end of preceding nucleotide (60 nucleotides/sec) the non-coding strand of DNA reforms a DNA double helix by pairing with the coding strand
12 3. TERMINATION transcription proceeds until RNA polymerases reaches a TERMINATOR site on the DNA; RNA molecule is then released Segment of DNA transcribed into one RNA = TRANSCRIPTION UNIT
13 Eukaryotic genes have junk! Eukaryotic genes are not continuous exons = the real gene expressed / coding DNA Exit the nucleus introns stay in the nucleus! introns = the junk inbetween sequence Stay in the nucleus eukaryotic DNA intron = noncoding (inbetween) sequence exon = coding (expressed) sequence
14 mrna splicing Post-transcriptional processing eukaryotic DNA eukaryotic mrna needs work after transcription primary transcript = pre-mrna mrna splicing edit out introns make mature mrna transcript intron = noncoding (inbetween) sequence ~10,000 bases primary mrna transcript mature mrna transcript exon = coding (expressed) sequence pre-mrna ~1,000 bases spliced mrna
15 RNA splicing enzymes snrnps small nuclear RNA proteins Spliceosome several snrnps recognize splice site sequence cut & paste gene exon No, 5' not smurfs! snurps exon mature mrna 5' snrna intron snrnps exon 5' 3' 5' exon 3' spliceosome 3' lariat 3' excised intron
16 More post-transcriptional processing Need to protect mrna on its trip from nucleus to cytoplasm enzymes in cytoplasm attack mrna protect the ends of the molecule add 5 GTP cap - METHYLATED GUANINE added to 5 end; for stability; prevents degradation used to bind mrna to ribosome add poly-a tail - stability; helps passage through nuclear membrane longer tail, mrna lasts longer: produces more protein 3' mrna A 5' G P P P
17 From gene to protein DNA transcription nucleus cytoplasm mrna translation ribosome a a a a a a a a a a a a a a a a a a a a a a protein trait
18 Translation from nucleic acid language to amino acid language Occurs on RIBOSOMES in CYTOPLASM in both PROKARYOTES & EUKARYOTES
19 How does mrna code for proteins? 4 DNA ATCG TACGCACATTTACGTACGCGG mrna 4 protein 20 AUCG AUGCGUGUAAAUGCAUGCGCC? Met Arg Val Asn Ala Cys Ala How can you code for 20 amino acids with only 4 nucleotide bases (A,U,G,C)?
20 mrna codes for proteins in triplets DNA mrna TACGCACATTTACGTACGCGG codon AUGCGUGUAAAUGCAUGCGCC GUA AAU GCA GCC protein? Met Arg Val Asn Ala Cys Ala
21 Cracking the code Crick - determined 3-letter (triplet) codon system WHYDIDTHEREDBATEATTHEFATRAT MESSENGER RNA carries DNA message from nucleus to cytoplasm; message is read in triplets called CODONS 64 different codons code for 20 different amino acids; 3 bases = 1 codon = 1 amino acid
22 The code Code for ALL life! strongest support for a common origin for all life Code is redundant several codons for each amino acid 3rd base wobble - codons for same amino acid can differ in 3 rd base Why is the wobble good? Start codon AUG methionine Stop codons UGA, UAA, UAG
23 How are the codons matched to amino acids? DNA 3 TACGCACATTTACGTACGCGG 5 mrna trna amino acid 5 3 AUGCGUGUAAAUGCAUGCGCC UAC Met 5 GCA Arg CAU Val codon anti-codon 3
24 Transfer RNA structure Clover leaf structure anticodon on clover leaf end amino acid attached on 3 end
25 Loading trna Aminoacyl trna synthetase enzyme which bonds amino acid to trna bond requires energy ATP AMP bond is unstable so it can release amino acid at ribosome easily activating enzyme Trp C=O Trp C=O Trp OH H 2 O OH O O trna Trp anticodon tryptophan attached to trna Trp A C C U G G mrna trna Trp binds to UGG condon of mrna
26 Ribosomes Ribosomes not making proteins exist as separate subunits Ribosomes making proteins for mebranes/ export: proteins are tagged so can be attached to rough ER; Cytoplasmic proteins made on free ribosomes
27 Ribosomes Facilitate coupling of trna anticodon to mrna codon Structure ribosomal RNA (rrna) & proteins 2 subunits: large + small Prokaryotic and eukaryotic subunits are different sizes = evidence for Endosymbiotic theory PROKARYOTIC RIBOSOMES: 30S + 50S = 70S; EUKARYOTIC RIBOSOMES: 40S + 60S = 80S E P A
28 Ribosomes A site (aminoacyl-trna site) holds trna carrying next amino acid to be added to chain P site (peptidyl-trna site) holds trna carrying growing polypeptide chain E site (exit site) empty trna leaves ribosome from exit site 5' E U A C A U G P Met A 3'
29 Building a Protein 1. Initiation Small ribosomal subunit attaches to the 5' end of the mrna ('start' codon - AUG) energy comes from GTP (guanosine triphosphate) trna carries 1 st amino acid (METHIONINE) to the mrna large ribosomal subunit attaches to the mrna
30 2. Elongation Ribosome moves along mrna matching trna ANTICODONS with mrna CODONS trna with new amino acid attaches at A site trna at A site moves to P site and receives growing chain trna a P site moves to E site and exits Released trna can recycle and bring in a new amino acid a new trna enters the A site and repeats the process increasing the polypeptide chain Leu length Met Met trna Met Met Leu Leu Leu Val Ser Ala Trp release factor UAC GAC 5' UAC 5' UACGAC AA C AAU 5' AUG C UGAAU 5' mrna A UG C UG U AUG UG 3' 3' 3' E P A UAC GAC C AA U AUG UG 3' A CC U GG UA A 3'
31 3. TERMINATION occurs when the ribosome encounters a 'stop' codon ribosome subunits detach; polypeptide is released mrna can be reread multiple time POLYSOMES- = strings of ribosomes can work on same mrna at same time
32 Protein targeting Signal peptide address label start of a secretory pathway Destinations: secretion nucleus mitochondria chloroplasts cell membrane cytoplasm etc
33 SIGNAL-RECOGNITION PARTICLE (SRP) Protein synthesis begins on free ribosomes Polypeptides that will become MEMBRANE PROTEINS or be SECRETED are marked SRP (SIGNAL RECOGNITION PARTICLE) attaches to protein signal sequence and receptor on ER Growing protein chain is inserted into ER lumen, Complex disconnects
34 SRP
35 POST TRANSLATIONAL MODIFICATION Changes to polypeptide chain to make it a protein CHAPARONINS-help wrap into 3D shape Some have groups added (sugars, lipids, phosphates, etc) EX: glycoproteins (protein + sugar) Some have segments removed ZYMOGEN (OR proenzyme) = inactive enzyme precursor Requires a biochemical change to become an active enzyme Usually occurs in LYSOSOMES where specific part is cleaved EX: insulin made as one chain middle removed to become active
36 Prokaryote vs. Eukaryote genes Prokaryotes eukaryotic DNA DNA in cytoplasm circular chromosome naked DNA (Eubacteria only) no introns exon = coding (expressed) sequence Eukaryotes DNA in nucleus linear chromosomes DNA wound on histone proteins introns and exons intron = noncoding (inbetween) sequence
37 Translation in Prokaryotes Transcription & translation are simultaneous in bacteria DNA is in cytoplasm no mrna editing ribosomes read mrna as it is being transcribed
38 DNA RNA polymerase Can you tell the story? pre-mrna exon intron 5' GTP cap amino acids trna large ribosomal subunit mature mrna poly-a tail polypeptide aminoacyl trna synthetase 3' 5' small ribosomal subunit E P A trna ribosome
39 The Transcriptional unit (gene?) enhancer b 20-30b translation start exons translation stop 3' RNA polymerase TATA TAC transcriptional unit (gene) ACT 5' DNA DNA UTR introns UTR promoter transcription start transcription stop 5' 3' pre-mrna 5' 3' GTP mature mrna AAAAAAAA
40 Bacterial chromosome Protein Synthesis in Prokaryotes Transcription mrna Psssst no nucleus! Cell membrane Cell wall
41 Any Questions?? What color would a smurf turn if he held his breath?
42 Substitute Slides for Student Print version
43
44 The Transcriptional unit enhancer b 20-30b exons 3' RNA polymerase TATA TAC transcriptional unit ACT 5' DNA introns 5' 3' 5' 3'
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