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Chapter 10 Gene Action and Expression Central Dogma DNA is the genetic material within the nucleus. Replication The process of replication creates new copies of DNA. The process of transcription creates an RNA using DNA information. DNA Transcription RNA Nucleus The process of translation creates a protein using RNA information. Translation Protein Cytoplasm Two types of nucleic acids # of strands kind of sugar bases used Two types of nucleic acids RNA DNA • Carries proteinencoding information • Carries RNAencoding information • Can be catalytic • Not catalytic Types of RNA mRNA Messenger RNA encodes protein rRNA Ribosomal RNA part of ribosome,used to translate mRNA into protein tRNA Transfer RNA couples the region which binds the mRNA codon and its amino acid rRNA is part of ribosome, used to translate mRNA into protein tRNA is a connection between anticodon and amino acid The genetic code • There is a 3 to 1 correspondence between RNA nucleotides and amino acids. • The three nucleotides used to encode one amino acid are called a codon. • The genetic code refers to which codons encode which amino acids. The genetic code A codon of three nucleotides determines choice of amino acid The genetic code is nonoverlapping The genetic code is universal • All known organisms use the same genetic code. • (Rare organisms use one codon for an additional amino acid.) The genetic code is degenerate Some codons encode the same amino acid. e.g. GGU, GGC, GGA, and GGG all encode glycine Degeneracy is mostly at the third base of the codon. Some codons have additional functions AUG encodes methionine. Methionine can be used within a protein sequence and is often the first amino acid cueing the beginning of translation. UAA, UAG, and UGA do not encode an amino acid These codons signal termination of the protein. Transcription • Three step process: Initiation Control of transcription is regulated by transcription factors. Elongation RNA polymerase adds nucleotides to growing RNA. Termination Sequences in the DNA prompt the RNA polymerase to fall off ending the transcript. Initiation of transcription Transcription begins at regions called the promoters. The promoter recruits TATA protein, a DNA binding protein, which in turn recruits other proteins. TATA binding protein Promoter DNA GG Transcription factor Gene sequence to be transcribed TATA CCC TATA box Transcription begins When a complete transcription complex is formed RNA polymerase binds and transcription begins. • Transcription DNA is used as a template for creation of RNA using • the enzyme RNA polymerase. DNA 5 ’ G T C A T T C G G 3’ 3’ C A G T A A G C C 5’ Transcription • RNA polymerase reads the nucleotides on the template strand from 3’ to 5’ and creates an RNA molecule that looks like the coding strand. DNA DNA coding strand 5 ’ G T C A T T C G G 3’ 3’ C A G T A A G C C DNA template strand 5’ Transcription DNA coding strand 5 ’ DNA G T C A T T C G G 3’ 3’ G U C A U U C G G 3’ C A G T A A G C C 5’ DNA template strand 5 ’ RNA mRNA processing • mRNA transcripts are modified before use as a • template for translation: •Addition of capping nucleotide at the 5’ end •Addition of polyA tail to 3’ end Important for moving transcript out of nucleus And for regulating when translation occurs •Splicing occurs removing internal sequences introns are sequences removed exons are sequences remaining mRNA processing • Gene Expression • Together transcription and translation are called gene expression. • The genetic information encoded in the DNA of an embryo includes all of the genes needed to develop and maintain the organism. • Different cell types express different subsets of genes. • Differential gene expression during development establishes the role of a cell within the body. Translation • The process of reading the RNA sequence of an mRNA and creating the amino acid sequence of a protein is called translation. DNA Transcription T T C A G T C A G A A G U C A G U C DNA template strand Messenger RNA mRNA Codon Codon Codon Translation Protein Lysine Serine Valine Polypeptide (amino acid sequence) Translation is composed of three steps Initiation translation begins at start codon (AUG=methionine) Elongation the ribosome uses the tRNA anticodon to match codons to amino acids and adds those amino acids to the growing peptide chain Termination translation ends at the stop codon UAA, UAG or UGA Translation initiation Initiation Complex Small ribosomal subunit 5’ 3’ Leader mRNA sequence mRNA U U C G U C A U G G G A U G U A A G C G A A U A C Initiator tRNA Assembling to begin translation Met tRNA of methione hydrogen bonds with the anticodons UAC at the start codon AUG on the mRNA small rRNA subunit Translation Elongation Ribosome 5’ 3’ mRNA A U G G G A U G U A A G C G A U A C C C U tRNA Amino acid Met Gly Large ribosomal subunit Translation Elongation • Large rRNA subunit attaches to the small rRNA subunit • tRNA of the second amino acid complementary to the second triplet code attaches by H bonds • Enzyme catalyzes the dehydration synthesis reaction between the two amino acids • MettRNA is released and leaves the complex to go back and get recharged with more met. Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A U A C C C U Met Gly Translation Elongation • The rRNA complex moves one triplet space • The third charged tRNA attaches by the anticodons to the third triplet codon on mRNA • These steps are repeated and amino acids are bonded to make the protein sequence Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A C C U A C A Gly Cys Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A C C U A C A Gly Cys Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A A C A U U C Cys Lengthening polypeptide (amino acid chain) Lys Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A A C A U U C Cys Lys Translation Elongation 5’ 3’ mRNA A U G G G A U G U A A G C G A A C A U U C Cys Lys Translation Elongation Stop codon 5’ mRNA A U G G G A U G U A A G C G A U A A U U C Lys Release factor Translation Termination Stop codon Ribosome reaches stop codon 5’ mRNA A U G G G A U G U A A G C G A U A A Release factor Translation Termination Once stop codon is reached, elements disassemble. Release factor Translation Termination • Elongation continues till the stop codon is reached • A release factor attaches to the stop codon • The rRNAs and mRNA complex is broken • Protein released Translation: multiple copies of a protein are made simultaneously Levels of protein structure sequence of amino acids Primary structure Secondary structure shapes formed with regions of the protein (helices, coil, sheets) Tertiary structure shape of entire folded protein due to interactions between particular peptides Quaternary structure structures formed by interaction of several proteins together e.g. Functional hemoglobin is two alpha-hemoglobin proteins and two beta-hemoglobin proteins form a heterotetramer Levels of protein structure Misfolding of protein impairs function •Misfolded prion protein disrupts functions of other normally folded prion proteins. •Aberrant conformation can passed on propagating like an “infectious” agent. Human Genome • 3.2 million DNA base pairs • 1.5% encode proteins < = > 98.5% not protein encoding • ~ 31,000 genes encoding 100,000 - 200,000 proteins • How are 100,000 to 200,000 proteins produced from 31,000 genes? • What is the 98.5% of the human genome that does not encode proteins? Alternative splicing of exons forms distinct proteins: one gene, many proteins Exon shuffling forms distinct proteins: several genes, multiple proteins Noncoding portion of the human genome Type of sequence Function or characteristic Noncoding RNAs Translation (tRNA,rRNA) Pseudogenes RNA processing Introns Removed with RNA processing Promoters and other regulatory regions Determine when and where transcription occurs Repeats: Transposons DNA that moves around genome Telomeres Chromosome tips Centromeres Important for attachment to spindle Duplications Unknown Simple short repeats unknown Transposons • Mobile DNA elements • First identified in corn by Barbara McClintock in 1940s • “selfish DNA”? • Grouped by size and sequence similiarity: • LINES • • long interspersed elements of 6,000 bases transcribed and trimmed to 900 bases SINES short interspersed elements of 100-500 bases • Alu element most common SINE in humans • 300,000 to 500,000 copies => 2-3% of genome! • Gene expression changes during development