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CHMI 2227E Biochemistry I

CHMI 2227E Biochemistry I. Gene expression. Nature of the gene. Genes are discrete regions of a DNA molecule Genes are permanent fixtures of your DNA; Every single cell in your body (every 100 trillion [i.e. 10 -15 cells] of them) has the exact, same DNA, the exact, same set of genes.

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CHMI 2227E Biochemistry I

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  1. CHMI 2227EBiochemistry I Gene expression CHMI 2227 - E.R. Gauthier, Ph.D.

  2. Nature of the gene • Genes are discrete regions of a DNA molecule • Genes are permanent fixtures of your DNA; • Every single cell in your body (every 100 trillion [i.e. 10-15 cells] of them) has the exact, same DNA, the exact, same set of genes. • Genes have « encoding » property: • The order of the nucleotide bases in a given gene (their sequence) has meaning; • For example: the order of nucleotide bases in the myoglobin gene has all the required information to make the myoglobin protein. CHMI 2227 - E.R. Gauthier, Ph.D.

  3. CHMI 2227 - E.R. Gauthier, Ph.D.

  4. Nature of the gene • But: chimps, humans and even worms have the SAME NUMBER OF GENES… • Moreover, most of these genes are SHARED between these three species… • What the %$#@? CHMI 2227 - E.R. Gauthier, Ph.D.

  5. Genes can be turned on or off (i.e. regulated): Temporally: e.g. during development; Spatially: e.g. turned on in the brain/ turned off in the liver. Timing and location (i.e. regulation) of gene expression is more important than the actual number of genes. Nature of the gene • Different homeotic (Hox) genes (colors) are turned on (expressed) at specific time during development and lead to the formation of specific body structures. CHMI 2227 - E.R. Gauthier, Ph.D.

  6. Normal fly Antennapedia mutation What happens when gene expression is screwed-up? • Homeotic genes are responsible for the development of discrete regions of the body: • Antp (antennapedia) specifies the formation of legs wings/thorax (flies) / thorax (humans) • Dfd (deformed) specifies the formation of the head (flies)/neck (humans) • If the Antp gene is expressed in the same cells as Dfd, legs will grow on the head of the fly. That’s what happens in flies bearing the Antennapedia mutation. CHMI 2227 - E.R. Gauthier, Ph.D.

  7. Central dogma of molecular biology CHMI 2227 - E.R. Gauthier, Ph.D.

  8. Transcription factors Promoter Transcription: turning on a gene • Each gene is preceded by a stretch of DNA called the promoter; • The promoter has DNA sequences that allow the recruitment of a distinct set of proteins called transcription factors; • These transcription factors help to recruit RNA polymerase, the enzyme which catalyses the synthesis of an RNA molecule complementary and antiparallel to one of the DNA strands of the gene (the template strand). CHMI 2227 - E.R. Gauthier, Ph.D.

  9. Promoters • Two major sequences are found in bacterial promoters: • TATA box (aka Pribnow box): • Centered 10 bp away from the transcription initiation site (denoted +1 below); • Directly binds RNA pol • Tells the RNA pol where to start transcription • -35 box: • Centered 35 bp away from the transcription initiation site • Directly binds RNA pol • Helps to stabilize the binding of RNA pol to the promoter CHMI 2227 - E.R. Gauthier, Ph.D.

  10. Promoters • Variations in the nucleotide sequence of the promoter are responsible for changes in the timing of gene expression; CHMI 2227 - E.R. Gauthier, Ph.D.

  11. Transcription CHMI 2227 - E.R. Gauthier, Ph.D.

  12. Transcription Messenger RNA (mRNA) is antiparallel to template strand Complementarity (5’ 3’) CHMI 2227 - E.R. Gauthier, Ph.D.

  13. mRNAs are modified in eukaryotes CHMI 2227 - E.R. Gauthier, Ph.D.

  14. In bacteria, the mRNA is nice and ready to make proteins (i.e. being translated) as soon as it is produced; In eukaryotes, the mRNA needs to be further processed before it can be translated: Removal of introns  splicing Addition of a non-coded GTP nucleotide at the 5’ end of the mRNA  capping Addition of 50-200 non-coded adenosine residues at the 3’end of the mRNA  polyadenylation Export of the mRNA out of the nucleus and into the cytoplasm (because translation ONLY occurs in the cytoplasm). mRNAs are modified in eukaryotes Introns: part of the gene which is transcribed but NOT found in the mRNA. Exons: part of the gene which ends up in the mRNA after transcription. CHMI 2227 - E.R. Gauthier, Ph.D.

  15. mRNAs are translated into proteins • During translation, the nucleotide sequence of the mRNA is read and decoded into an amino acid sequence. CHMI 2227 - E.R. Gauthier, Ph.D.

  16. Translation • To translate an mRNA into a protein, the following ingredients are needed: • mRNA template • Amino acids • Transfer RNA (tRNA): • adaptor between amino acid and mRNA • In charge of converting the nucleotide sequence code into an amino acid sequence. • Ribosomes: • organelles directing the translation process. CHMI 2227 - E.R. Gauthier, Ph.D.

  17. Short RNA molecule (between 73 and 95 nucleotides long); Has two major features: The acceptor arm: where a specific amino acid is covalently coupled; The anticodon arm: has a 3-nucleotide sequence: the anticodon The anticodon base pairs with a complementary and antiparallel, 3 nucleotide sequence on the mRNA: the codon; The genetic code is the relationship between the sequence of a codon with a specific amino acid. Transfer RNA (tRNA) CHMI 2227 - E.R. Gauthier, Ph.D.

  18. 3’ 5’ 5’ 3’ A U mRNA The genetic code • Each tRNA is bound to a unique, specific amino acid; • The nature of this amino acid depends on the sequence of the anticodon; • Since the anticodon on the tRNA base pairs with the codon on the mRNA, there is a relationship between the sequence of the codon on the mRNA and the amino acid bound to the tRNA: this relationship is the GENETIC CODE. • Thus, in this example, the mRNA codon 5’UGU3’ codes for the amino acid Cys. • The genetic code is (almost) universal: the meaning of each codon is (pretty much) the same whatever organism is studied. CHMI 2227 - E.R. Gauthier, Ph.D.

  19. The genetic code CHMI 2227 - E.R. Gauthier, Ph.D.

  20. Translation • Translation is the phenomenon whereby the mRNA code (i.e. the sequence of codons) is decoded into an amino acid sequence; • Translation involves three major steps: • Initiation: recognition of the translation starting point by the ribosome and a special tRNA; • Elongation: • successive binding of tRNAs to the ribosome; • formation of a peptide bond between an incoming amino acid and the growing polypeptide chain; • translocation of the ribosome to decode the next codon; • Termination: • One of three « stop » codons reaches the ribosome • The ribosome is dissociated, freeing the mRNA and the completed polypeptide; CHMI 2227 - E.R. Gauthier, Ph.D.

  21. fMET = N-formyl methionine The initiation codon is ALWAYS AUG  Methionine Translation - Initiation http://biology.unm.edu/ccouncil/Biology_124/Images/RNAtranslation.jpeg CHMI 2227 - E.R. Gauthier, Ph.D.

  22. Peptide bond: synthesized by the « peptidyl transferase » enzyme on the large ribosomal subunit Translocation Translation - Elongation http://biology.unm.edu/ccouncil/Biology_124/Images/RNAtranslation.jpeg CHMI 2227 - E.R. Gauthier, Ph.D.

  23. Three « Stop » codons exist: UGA; UAA, UAG Translation - Termination http://biology.unm.edu/ccouncil/Biology_124/Images/RNAtranslation.jpeg CHMI 2227 - E.R. Gauthier, Ph.D.

  24. Cue from environment (hormones, temperature, etc) Protein folding Protein function Change in cell behavior Change in organism Adaptation to environment http://www.contexo.info/DNA_Basics/images/gene_expression.gif CHMI 2227 - E.R. Gauthier, Ph.D.

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