No growth: Mutant cells cannot grow and divide. Classes of Neurospora crassa. Class I mutants Class II mutants Class III mutants

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1 XPRIMNT Growth: Wild-type cells growing and dividing Minimal medium No growth: Mutant cells cannot grow and divide RSULTS Condition Minimal medium (MM) (control) MM + ornithine MM + citrulline Wild type Classes of Neurospora crassa Class I mutants Class II mutants Class III mutants MM + arginine (control) CONCLUSION Wild type Class I mutants (mutation in gene A) Class II mutants (mutation in gene B) Class III mutants (mutation in gene C) Gene A Gene B Gene C Precursor Precursor Precursor Precursor nzyme A nzyme A nzyme A nzyme A Ornithine Ornithine Ornithine Ornithine nzyme B nzyme B nzyme B nzyme B Citrulline Citrulline Citrulline Citrulline nzyme C nzyme C nzyme C nzyme C Arginine Arginine Arginine Arginine 1 TRANSCRIPTION TRANSLATION Ribosome Polypeptide (a) Bacterial cell Nuclear envelope TRANSCRIPTION PROCSSING Pre- TRANSLATION Ribosome Polypeptide (b) ukaryotic cell 2

2 molecule Gene 1 Gene 2 Gene 3 template strand TRANSCRIPTION Codon TRANSLATION Protein Amino acid 3 Second base First base ( end of codon) Third base ( end of codon) 4

3 (a) Tobacco plant expressing a firefly gene (b) Pig expressing a jellyfish gene 5 Promoter Transcription unit Start point polymerase 1 Initiation longation Nontemplate strand of Unwound transcript Template strand of 2 longation polymerase end nucleotides Rewound transcript 3 Termination Completed transcript Newly made Direction of transcription ( downstream ) Template strand of 6

4 1 A eukaryotic promoter includes a TATA box Promoter Template TATA box Start point Template strand Transcription factors 2 Several transcription factors must bind to the before polymerase II can do so. polymerase II 3 Additional transcription factors bind to the along with polymerase II, forming the transcription initiation complex. Transcription factors transcript Transcription initiation complex 7 G P Protein-coding segment Polyadenylation signal P P AAUAAA AAA AAA Cap UTR Start codon Stop codon UTR Poly-A tail 8

5 Pre- Cap xon Intron xon Intron xon 146 Poly-A tail Coding segment Introns cut out and exons spliced together Cap Poly-A tail 1 UTR 146 UTR 9 transcript (pre-) xon 1 Intron xon 2 Protein sn snrnps Other proteins Spliceosome Spliceosome components xon 1 xon 2 Cut-out intron 10

6 Gene xon 1 Intron xon 2 Intron xon 3 Transcription processing Translation Domain 3 Domain 2 Domain 1 Polypeptide 11 Polypeptide Amino acids Ribosome t with amino acid attached Phe Gly t Anticodon Codons 12

7 Amino acid attachment site Hydrogen bonds Anticodon (a) Two-dimensional structure Amino acid attachment site Hydrogen bonds Anticodon (b) Three-dimensional structure Anticodon (c) Symbol used in this book 13 Amino acid Aminoacyl-t synthetase (enzyme) P P P ATP Adenosine P i P Adenosine P P i P i t Aminoacyl-t synthetase t P Adenosine AMP Computer model Aminoacyl-t ( charged t ) 14

8 t molecules Growing polypeptide P A xit tunnel Large subunit Small subunit (a) Computer model of functioning ribosome P site (Peptidyl-t binding site) site (xit site) binding site P A A site (Aminoacylt binding site) Large subunit Small subunit (b) Schematic model showing binding sites Amino end Growing polypeptide Next amino acid to be added to polypeptide chain t Codons (c) Schematic model with and t 15 Met U A A U C G P site Met Large ribosomal subunit Initiator t Start codon binding site Small ribosomal subunit GTP GDP A Translation initiation complex 16

9 Amino end of polypeptide Ribosome ready for next aminoacyl t P A site site GTP GDP P A P A GDP GTP P A 17 Release factor Free polypeptide Stop codon (UAG, UAA, or UGA) 2 GTP 2 GDP 18

10 Growing polypeptides Completed polypeptide Incoming ribosomal subunits (a) Start of ( end) Polyribosome nd of ( end) Ribosomes (b) 0.1 µm 19 Ribosome Signal peptide Signalrecognition particle (SRP) CYTOSOL R LUMN SRP receptor protein Translocation complex Signal peptide removed R membrane Protein 20

11 Wild-type hemoglobin C T T G A A Mutant hemoglobin C A T G T A G A A G U A Normal hemoglobin Glu Sickle-cell hemoglobin Val 21 template strand Protein Amino end Wild-type Stop Carboxyl end A instead of G U instead of C Stop Silent (no effect on amino acid sequence) xtra A xtra U Stop Frameshift causing immediate nonsense (1 base-pair insertion) T instead of C missing A instead of G missing Stop Missense Frameshift causing extensive missense (1 base-pair deletion) A instead of T missing U instead of A missing Stop Stop Nonsense No frameshift, but one amino acid missing (3 base-pair deletion) (a) Base-pair substitution (b) Base-pair insertion or deletion 22

12 polymerase Polyribosome polymerase Direction of transcription 0.25 µm Polyribosome Polypeptide (amino end) Ribosome ( end) 23 TRANSCRIPTION transcript PROCSSING xon polymerase transcript (pre-) Intron NUCLUS Poly-A Aminoacyl-t synthetase CYTOPLASM Amino acid t AMINO ACID ACTIVATION A P Ribosomal subunits Cap Growing polypeptide Activated amino acid Poly-A Cap TRANSLATION A Anticodon Codon Ribosome 24