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Copying of the DNA. Such as when bacteria divide they copy their single circular chromosome so that each daughter cell will have a single copy. This is done by enzymes in the cell called DNA polymerases |
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Making an RNA copy of a gene. Messenger RNA. This carries the information in the DNA to the cell. The enzyme that does this is called an RNA polymerase. |
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Converting the mRNA into protein by reading the three letters at a time and hooking together the correct amino acids to make a protein. Ribosomes do this. |
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The way that information is stored in DNA is in the sequence of the nucleotides taken 3 at a time. Each 3 DNA nucleotides in a gene code for one of the 20 amino acids. Remember that the DNA is transcribed into mRNA and it is the mRNA code that is actually read by ribosomes to make protein. |
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Three letter sequence that codes for an amino acid. |
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Not all genes are active at the same time. If they were it would constitute a tremendous waste of energy. Instead they are turned on (transcribed) only when they are needed. |
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Genes that are not on all of the time are called inducible and they can be turned on by inducers- molecules that turn on transcription |
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Genes that are always turned on (transcribed) are called |
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Genes that are always turned on (transcribed) are called constitutive. |
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Central Dogma in Prokaryotes |
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A. Prokaryote -No nucleus -the genes are not interrupted by non coding regions -several adjacent genes are often transcribed at the same time as one large mRNA. -These are translated into separate proteins. |
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Because of the bond angles formed by hydrogen bonding of the bases, the two strands of antiparallel DNA form a right-handed helix. |
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major groove; minor groove |
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The two sugar-phosphate backbones form ridges in the surface of the DNA molecule. The space between the ridges are grooves. One groove is small and is called the Minor Groove. The other is bigger and is called the Major Groove. |
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DNA absorbs light at a wavelength of _____nm more than any other wavelength |
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Single-stranded DNA (ssDNA) absorbs more UV light than double-stranded DNA (dsDNA) so when dsDNA melts (denatures) you get an increase in absorbance |
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Melting Temperature: Temperature at which half of the hydrogen bonds between the two strands are broken. ~85 |
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In prokaryotes, proteins organize the chromosome into supercoiled domains Involved in gene regulation |
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allows the DNA to be packed into the cell. |
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when the DNA is twisted in the opposite direction of the helix and positive supercoiling is when the DNA is twisted in the same direction as the helical turns. NEGATIVE supercoiling is most common in nature. Gyrase (a form of topoisomerase) is the enzyme that does this. |
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have been discovered in extreme thermophiles This may stabilize their DNA in hot environments. Reverse gyrase is an enzyme that catalyzes positive supercoiling in some extreme thermophile |
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is an enzyme that cleaves the phosphodiester bond in a single strand in the DNA and causes the tension to relax. This removes the supercoiling. |
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topoisomerase II or DNA gyrase |
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catalyzes negative supercoiling of DNA. This enzyme is very important to the survival of the cell. |
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antibiotics that inhibit DNA gyrase and kill bacterial cells. |
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Naladaxic acid, novobiocin, and ciprofloxacin |
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important in determining when genes are "turned on" and "turned off" |
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sequences can be inverted and read the same way; create the possibility for interesting structures in DNA like hairpins and cruciform structures |
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nucleic acids that: 1. Have genes. 2. Replicate themselves. Chromosome Plasmid Virus Mitochondrion, chloroplast (eukaryotes only) Transposable elements-can move from one place to another Transposons-genes with transposable elements on either side-jumping gene |
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Nonchromosomal genetic elements |
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A. Viruses and viroids;B. Plasmids:; C. Mitochondria and chloroplasts; D. Transposable elements (Transposons) |
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only thing that is passed on to the next generation in all organisms is |
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DNA (and in most eukaryotes-mitochondria from your mother) |
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With a single origin we get "Theta replication" because the DNA looks like the Greek letter Theta (Circular DNA replication) |
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A special RNA polymerase called Primase adds a short piece (11 ribonucleotides) of RNA to newly exposed DNA strands. |
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adds deoxynucleotides to the RNA primer using its free 3’-OH |
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small pieces of the lagging strand |
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In the lagging strand, DNA polymerase III eventually runs into the primer from an adjacent replicating site. When this happens, DNA polymerase I digests the RNA and then fills in the gap using the 3’ end left by polymerase III. |
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Requirements for DNA replication |
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Definition
1. Template: 2. Primer 3. Primase 4. DNA polymerase III: 5. Deoxynucleotide triphosphates 6. DNA polymerase I: 7. DNA helicase 8. Single-stranded binding protein 9. DNA Ligase: |
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must have something to copy. The strand that is copied is called the template. |
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Must have a starting place. You must have something to hook the nucleotides to as they are brought in place. This is a short strand of RNA. Remember that the nucleotides are added in a 5' to 3' direction. |
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Deoxynucleotide triphosphates |
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Definition
dATP, dTTP, dCTP, dGTP. Precursors for the growing chain. |
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When replication is well under way this enzyme chews up the RNA primer and replaces it with DNA. |
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This is a DNA unwinding protein. |
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Single-stranded binding protein |
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Proteins that bind to the unwound (ss) DNA and prevent it from re base pairing. |
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Attaches the ends of two DNA strands. |
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Begins at the origin of replication. A. Origin binding protein (DnaA) binds to the origin and then Helicase binds and separates the double strands. B. Single stranded binding proteins bind and prevent the strands from repairing. C. Primase makes a short piece of RNA on the leading strand and DNA polymerase III starts synthesizing DNA. D. Primase makes an RNA primer in 5' to 3' direction of the lagging strand. E. Replication begins on the two strands forming the replication fork |
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A. DNA Polymerase III adds DNA to the primer in the 5' to 3' direction. B. DNA Polymerase I degrades the RNA primer and replaces it with DNA. |
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Termination (Replication) |
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Definition
A. The polymerases converge on a region of the DNA containing sequences known as ter sites where they meet and complete the replication process. B. The single DNA strands are joined by DNA ligase |
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A complex of enzymes involved in replicating DNA |
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The error rate in DNA replication is very low: 1 in a billion to 1 in a trillion. This is accomplished in what 3 ways? |
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1. Base pairing; 2. proofreading; 3. Mismatch repair |
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DNA polymerase III reads the template nucleotides accurately and usually inserts the correct complementary nucleotide. |
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DNA polymerase III is able to detect incorrect nucleotides that are not complementary and remove them by chewing back the new strand from the 3' end (3' exonuclease activity). All prokaryotes, eukaryotes and viruses have this function. |
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by endouncleolytic proteins that are able to detect bases that are mispaired such as A-A or A-C. These enzymes work on DNA that has already been replicated long after the polymerase has gone on it's way. They excise the mismatched base and replace it with another. This is a type of endonuclease since it attacks DNA in the middle of a strand and not at the ends. |
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RNA mediated events in the cell: |
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Transcription and Translation |
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Transcription enzyme and product: |
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The enzyme is protein and the product is mRNA |
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Translation enzyme and product: |
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The enzyme is RNA and the product is protein |
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Three main types of RNA in the cell: |
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1. mRNA; messenger RNA. 2. tRNA; transfer RNA carries the amino acids-adaptor molecule. 3. rRNA; ribosomal RNA-Ribosomes are made of RNA and Protein and the RNA is called rRNA. |
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Transcription is catalyzed by: |
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RNAP is a ____-directed ____ polymerase |
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Name the 5 peptide subunits of RNAP |
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Definition
α2ββ'+σ + ω; α2ββ' is called the core enzyme; The main function of the sigma subunit is to find the proper place for the core enzyme to begin. |
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unit of genetic expression consisting of one or more related genes and the operator and promoter sequences that regulate their transcription |
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Who characterized the operon? |
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Originally characterized by Jacob and Monod in 1961 |
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Initiation:; Elongation; Termination: |
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Initiation (transcription) |
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Sigma finds the PROMOTER (a transcription on/off switch) and the core enzyme binds to the DNA and begins transcribing (making an RNA copy). |
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What is Sigma's function in transcription? |
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aids in recognition of promotor and initiation site. |
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Sigma dissociates and the core enzyme moves down the DNA, separating the strands just long enough to allow base pairing between the ribonucleoside triphosphates (ATP, UTP, CTP, GTP). Not dATP etc. Only one strand is read by the polymerase (coding strand) |
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The polymerase reaches a sequence in the mRNA called the terminator and falls off the DNA. |
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What are the Two types of transcription termination? |
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1. Rho dependent; 2. Rho independent (intrinsic); |
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Rho is a protein that travels behind the RNA Polymerase ON THE NEWLY MADE RNA and if the polymerase slows down the Rho protein catches up to it and causes it to fall off the DNA. Core slows down when it encounters certain termination sequences in the DNA called Rho utilization (rut) sequences. |
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Rho independent (intrinsic): |
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Definition
Inverted repeats form large hairpin structures in the newly made RNA and when these structures are followed by a string of uracils.
When the hairpin is formed the RNA polymerase slows down. Because the polyU strand is only weakly paired with the template strand, it dissociates from the template leaving the polymerase no substrate to add new nucleotides to thereby ending transcription |
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antibiotics that bind to and inhibit the beta subunit of RNA polymerase. |
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Rifamycins (Rifampicin, rifaldazine etc |
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they last a lot longer than mRNA does in the cell. tRNA, and rRNA are examples |
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1. aa-tRNA; 2. mRNA; 3. Ribosomes; 4. Factors |
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carries the genetic instructions from the DNA |
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catalyze assembly of the amino acids into peptides |
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proteins that are not part of the ribosome but are required for proper protein synthesis. |
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What structure does the tRNA have? |
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Who were awarded the Nobel Prize for the elucidation of the Genetic Code? |
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In 1968 Marshall Nirenberg, Gobind Khorana and Robert Holley |
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Bacterial ribosome composition |
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70S composed of 30S + 50S
30S composed of 21r + 16s; 50s composed of 34r + 23s + 5s |
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There are 3 tRNA binding sites on the ribosome: |
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Site where incoming aa-tRNA enters the ribosome. The tRNA is carried to the A-site as part of a ternary complex containing the aa-tRNA (1), a GTP molecule (2) and Elongation factor Tu (3). |
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Site where the tRNA containing the growing peptide chain is located. |
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Site where the P-site tRNA goes after it has given up its peptide to the tRNA in the A site. tRNA that has lost its amino acid is called deacylated tRNA. The tRNA is the E site is deacylated tRNA. The E-site tRNA exits the ribosome as the next aa-tRNA enters the A site to begin the next cycle. |
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Translation initiation; Formyl-Methionine-tRNA (Methionine with a formyl group on the end) |
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Definition
bound to the P-site during initiation and is therefore always the first amino acid in every protein in bacteria. |
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A sequence on the mRNA called the Shine- Dalgarno sequence is complementary to a sequence found in 16S RNA and these two sequences base pair and bind the mRNA to the ribosome |
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Definition
Amino acid (or peptide) is transferred to the tRNA in the A-site. Empty (deacylated)tRNA moves to the E-site (exit). A-site tRNA moves into the P-site and another aa-tRNA comes into the now empty A-site. |
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What proteins are required during elongation? |
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What protein does translocation require? |
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What proteins assist in termination? |
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You must have something in the ___ site to form the first peptide bond. |
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You must have something in the P site to form the first peptide bond. The fMet-tRNA specifically goes to the P-site first. All other tRNAs go to the A site. |
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Release factors recognize a stop codon in the A-site and the protein synthesis machinery disassembles |
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coupled transcription and translation. |
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Since there is no nucleus in prokaryotes, ribosomes can bind to the mRNA before it is finished being transcribed |
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In both eukaryotes and prokaryotes a single mRNA can have several ribosomes translating it at the same time and these mRNA/ribosome complexes are called polysomes. |
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Mistranslation rate of ribosomes in bacteria? |
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What determines the destination of the protein? |
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Protein does not contain signal sequence: non-secreted protein. Protein contains signal sequence: Protein inserted into membrane. Sec A: Protein secreted into periplasm |
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