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DNA vs Protein as genetic material
- DNA composed of 4 nucleotides, Protein composed of 20 aas
- DNA amount constant from cell to cell, protein amounts vary per cell
- DNA confined to nucleus, proteins throughout cell as enzymes
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characteristics of nucleic acids
- DNA composed of nucleotides, determined in 1800-1900s
- 3 parts of nucleotide: 5 carbon sugar, phosphate and nitrogenous base
- nucleotides held together by phosphodiester bonds
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Nucleotides
- composed of Adenine, guanine, cytosine and Uracil
- A 2 T
- C 3 G
- PAG, pure as gold
- pyrimidines are thymine, cytosine (or uracil)
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DNA vs RNA
- DNA double stranded
- DNA made of DEoxyribose sugar, phosphate group and nitrogenous base ATG or C
- RNA single stranded
- RNA made of RIBOSE sugar, phosphate group and nitrogenous base AUG or C
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1920's Frederick Griffith
showed component of virulent strain in Diplococcus could transform in non-virulent strains
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1940's Avery, MacLeod & McCarty
- Destruction of DNA in extracts of heat kills cells
- These extracts could NOT transform non-virulent cells
- RNA or protein destruction did not have same effect
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1950's Hershey and Chase
Confirmed NA heritable material: during phage replication in bacteria, parental DNA transferred to progeny, but NOT parental proteins
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Organic Chemistry
- Nitrogenous base attaches to C #1 of deoxyribose sugar
- Phosphate group attaches to C #5
- OH group of carbon 3 involved with phosphodiester bond
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Rosalind Franklin and Wilkins
- Through X Ray diffraction, showed:
- 1.DNA composed of 2 STRANDS of nucleotides, wound to form helix
- 2.both strands equally part throughout the molecule
- 3.helix diameter 20A and periodicities 3.4A and 34A
- Determined phosphate backbone on outside, with nitrogenous bases on inside
- Purine pairs w/Pyrimidine
- sugar phosphate backbone of each strain is antiparallel
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Genetic code triplet code
- degenerate: aa can be coded by more than 1 code
- unambiguous: each code encodes only 1 type of aa
- start (AUG) and stop signals (UAA, UGA, UAG)
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coding strand
- non-template strand of DNA
- IDENTICAL to the RNA that is transcribed
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template strand
- binds DNA and synthesizes complementary RNA strand
- only this strand is copied
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Transcription components
- RNA polymerase - composed of SIGMA subunit
- sigma subunit required for INITIATION, not elongation
- Elongation requires
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mRNA processing
- 5' 7-methyl cap in pro and euk
- only in euk 3' poly A tail (prevents degredation)
- only in euk, INTRONS spliced OUT
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2 mechanisms of splicing out INTRONS in eukaryotes
- 1. self-splicing, aka autocatalytic RNAs
- removes the introns from rRNA
- uses active site on intron to do transesterification reaction and release intron
- 2. splicesome
- RNA protein complex (snRNP) catalyzes removal of introns from mRNAs
- bind to active site, recruit more proteins and then cut b/w exon & intron, then fuses exons together
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translation process steps
- 1. initiation: initiation complex forms
- binding of small ribosomal subunit to mRNA
- binding of large ribosomal subunit to initiator tRNA
- 2. elongation
- reading of mRNA codons and adding of aa to polypeptide chain
- 3. termination
- release of polypeptide
- disassembly of ribosome
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Initiation in translation
- initiation factors 1,2 and 3 bind to small subunit of ribosome when mRNA binds
- IF3 released once initiator tRNA binds to mRNA codon in P site
- Large ribosomal subunit binds and IF1 and IF2 release
- EF-Tu binds to new tRNA now helping to allow entry into A site
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Elongation in translation
- fit of mRNA codon-anticodon tRNA checked by using energy from EF-Tu - GTP bond
- as more tRNAs attach to A site, PEPTIDYLtransferase (fxn of 28S rRNA) used to form peptide bonds
- mRNA shifts, through help from EF-G, allowing for A site to be open to allow another tRNA to enter
- New tRNA enters A site through help from EF-Tu and elongation continues
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Roles of peptidyl transferase in translation
- 1. forms peptide bonds between AAs from tRNA at P site and A site.
- 2. breaks high energy bonds between AAs and tRNA (this is the energy used for peptide bond)
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Termination in translation
- Release factor binds to A site when STOP codon is encountered
- These factors also aid in release of polypeptide chain from tRNA at P site
- RF1 binds to UAA, UAG
- RF2 binds to UAA, UGA
- Now Ribosome/mRNA complex disassembles
- Polypeptide now folds into its conformation, or assisted by chaperonins
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Euk vs prok translation
- Euk mRNA live for hrs, vs. prok mRNA which is minutes
- 7-methyl cap REQUIRED in euk
- Kozak recog. seq present in Euk
- majority of rRNA in euk is associated w/ ROUGH ER, vs. in prok where rRNA is in cytoplasm
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Protein folding bindings
- dependent on sequence of AAs
- Hydrogen bond formation of R group interaction of each other and environment
- Post-translational modification occurs, such as addition of phosphate, methyl, sugar, lipid group, formation of disulfide bonds, metal ions
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Post-translational modifications
- N-terminus AA removed or modified
- AA residues modified
- Carb. sidechains sometimes attached
- Polypeptide chains may be trimmed
- Signal seq. removed
- Polypeptide chains complexed w/metals
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Protein Structure
- 1. primary: aa sequence (polypeptide chain)
- 2. 2ndary: A helices and B pleated sheets are specific aas configured together closely
- 3. tertiary: 3D structure of fully folded protein
- 4. quaternary: 2 or more polypeptides forming the functional protein (ie: hemoglobin w/its 4 groups)
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