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Studying Genetics in the Twenty-First Century

CHAPTER

Genetics: The study of biological information

COURSE OUTLINE
1. Genes: Fundamental Informational Molecules of Life 2. Eukaryotic Gene regulation: Functional Molecules of Life Processes 3. Modern Genetic Techniques 4. Complex Systems and Molecular Interactions in cancer genetics

PART I

What Genes Are and What They Do

CHAPTER

Gene Expression: The Flow of Information from DNA to RNA to Protein

OUTLINE
1.1 The Genetic Code information to functional molecules 1.2 Transcription: From DNA to RNA 1.3 Translation: From mRNA to Protein 1.4 The Effect of Mutations on Gene Expression and Gene Function

Three levels of biological information


DNA Macromolecule made of nucleic acids Repository of the genetic code Proteins Macromolecules made of amino acids Amino acid sequence determined by DNA sequence Biological systems Network of interactions between molecules or groups of cells Accomplish coordinated functions
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Where and how is biological information stored


Biological function
= protein expression

=
gene expression

Appreciating the complexity of biological information


Double stranded DNA Hydrogen bonds Complementary base pairs composed of sugar, amine bonds, phosphate groups Conversion of a 3-D functional molecule to a 1-D molecule for computational analysis

Complementary base pairs are a key feature of the DNA molecule


DNA is comprised of four nitrogenous bases [guanine (G), adenine (A), cytosine (C), and thymine (T)], a deoxyribose, and a phosphate
G C and A T hydrogen bonds between each strand of the double helix

The two strands of the double helix are in opposite orientation

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Organization of genetic information in cells


Genes are sequences of DNA that encode proteins Chromosomes are organelles that package and manage the storage, duplication, expression, and evolution of DNA

Genomes are the entire collection of chromosomes in each cell of an organism


The human genome: 24 kinds of chromosomes 3 x 109 base pairs Encodes 20,000 30,000 genes
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Proteins are polymers of hundreds to thousands of amino acids


There are 20 different amino acids Information in DNA of a gene dictates the sequence of amino acids for the protein The order of amino acids determines the type of protein and its three dimensional structure Diversity of three-dimensional structure of protein generates an extraordinary diversity of protein function

The amino acid sequence determines the three-dimensional shape of the protein
Chemical formulas for two amino acids Three-dimensional shapes of two proteins

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RNA evolved into an intermediary in conversion of DNA information into protein


RNA is comprised of four nitrogenous bases [guanine (G), adenine (A), cytosine (C), and uracil (U)], a ribose, and a phosphate

Bases are read as triplets to encode amino acid subunits of protein

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All living organisms use essentially the same genetic code

Specific triplets of RNA bases encode the 20 amino acids

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Four general themes for gene expression


Pairing of complementary bases is the key to the transfer of information from DNA to RNA and from RNA to protein Polarities of DNA, RNA, and polypeptides help guide the mechanisms of gene expression Gene expression requires input of energy and participation of specific proteins and macromolecular assemblies Mutations that change genetic information or obstruct the flow of its expression can have dramatic effects on phenotype

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Mutation example in Predictive and Preventive Medicine


Genetic variations can cause or predispose
Single DNA base pair mutation in the betaglobin gene will lead to sickle cell anemia Mutation in the breast cancer BRCA1 gene results in a 70% chance in acquiring the disease

Differences: control of genetic expression

Gene expression: the flow of genetic information from DNA via RNA to protein
RNA polymerase transcribes DNA to produce an RNA transcript

Ribosomes translate the mRNA sequence to synthesize a polypeptide

Translation follows the "genetic code"


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Summary of the genetic code


Genetic code has triplet codons
Codons are nonoverlapping Three nonsense codons dont encode an amino acid; UAA (ocher), UAG (amber) and UGA (opal) Genetic code is degenerate Reading frame is established from a fixed starting point codon for translation initiation is AUG mRNAs and polypeptides have corresponding polarities Mutations can be created in three ways; frameshift, missense, and nonsense
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Transcription: From DNA to RNA


RNA polymerase catalyzes transcription

Promoters are DNA sequences that provide the signal to RNA polymerase for starting transcription
RNA polymerase adds nucleotides in 5-to-3 direction

Formation of phosphodiester bonds using ribonucleotide triphosphates (ATP, CTP, GTP, and UTP)
Hydrolysis of bonds in NTPs provides energy for transcription Terminators are RNA sequences that provide the signal to RNA polymerase for stopping transcription
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Structure of the methylated cap at the 5' end of eukaryotic mRNAs


Capping enzyme adds a "backward" G to the 1st nucleotide of a primary transcript

Transcribed bases

Methylated cap not transcribed

Triphosphate bridge

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Processing adds a tail to the 3' end of eukaryotic mRNAs

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RNA splicing removes introns

Exons sequences found in a genes DNA and mature mRNA (expressed regions)
Introns sequences found in DNA but not in mRNA (intervening regions) Some eukaryotic genes have many introns

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The human dystrophin gene: An extreme example of RNA splicing


Splicing removes introns from a primary transcript

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RNA processing splices out introns and joins adjacent exons


Short sequences in the primary transcript dictate where splicing occurs

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RNA processing splices out introns and joins adjacent exons (cont)
Two sequential cuts remove an intron

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Splicing is catalyzed by the spliceosome

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splicing video

http://www.youtube.com/watch?v=FVuAwBGw_pQ

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Translation: From mRNA to protein


Transfer RNAs (tRNAs) mediate translation of mRNA codons to amino acids Translation takes place on ribosomes that coordinate movement of tRNAs carrying specific amino acids

tRNAs are short single-stranded RNAs of 74 95 nt


Each tRNA has an anticodon that is complementary to an mRNA codon A specific tRNA is covalently coupled to a specific amino acid (charged tRNA) Base pairing between an mRNA codon and an anticodon of a charged tRNA directs amino acid incorporation into a growing polypeptide
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Some tRNAs contain modified bases

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Three levels of tRNA structure


Nucleotide sequence is the primary structure Secondary structure (cloverleaf shape) is formed because of short complementary sequences within the tRNA Tertiary structure (L shape) is formed by 3-dimensional folding
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Aminoacyl-tRNA synthetases catalyze attachment of amino acids to specific tRNAs


Each aminoacyl-tRNA synthetase recognizes a specific amino acid and the structural features of its corresponding tRNA

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Base pairing between an mRNA codon and a tRNA anticodon determines which amino acid is added to a growing polypeptide
Fig. 8.21

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Wobble: Some tRNAs recognize more than one codon for the amino acid they carry

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A ribosome has two subunits composed of RNA and protein

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Mechanism of translation
Initiation stage - start codon is AUG at 5 end of mRNA
In bacteria, initiator tRNA has formylated methionine (fMet)

Elongation stage - amino acids are added to growing polypeptide


Ribosomes move in 5-to-3 direction along mRNA 2-15 amino acids added to C terminus per second

Termination stage - polypeptide synthesis stops at the 3' end of the reading frame
Recognition of nonsense codons Polypeptide synthesis halted by release factors Release of ribosomes, polypeptide, and mRNA
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Different parts of a ribosome have different functions


Small subunit binds to mRNA Large subunit has peptidyl transferase activity Three distinct tRNA binding areas E, P, and A sites

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Translation of mRNAs on ribosomes: Initiation phase in prokaryotes


Ribosome binding site consists of a Shine-Dalgarno sequence and an AUG Three sequential steps: small ribosomal subunit binds first, fMet-tRNA positioned in P site, large subunit binds Initiation factors (not shown) play a transient role

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Translation of mRNAs on ribosomes: Initiation phase in eukaryotes


Small ribosomal subunit binds to 5' cap, then scans the mRNA for the first AUG codon Initiator tRNA carries Met (not fMet)

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Translation of mRNAs on ribosomes: Elongation phase


Addition of amino acids to C-terminus of polypeptide

Charged tRNAs ushered into A site by elongation factors (not shown)

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Polyribosomes consist of several ribosomes translating the same mRNA


Simultaneous synthesis of many copies of a polypeptide from a single mRNA

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Translation of mRNAs on ribosomes: Termination phase


No normal tRNAs carry anticodons for the stop codons

Release factors bind to the stop codons


Release of ribosomal subunits, mRNA, and polypeptide

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Posttranslational processing can modify polypeptide structure


(a) Cleavage may remove an amino acid (c) Chemical constituent addition may modify a protein

(b) Cleavage may split a polyprotein

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