Bio Exam 3

showed evidence of a 'transforming principle'

followed up on this to find that the 'transforming principle' was DNA

discovered the startling ratio of A:T and G:Cs

provided additional evidence that DNA was genetic material (at least in T2 bacteriophages!)

developed DNA double helix model based on others'evidence

unwinds the DNA

creates a short RNA fragment called a "primer"

synthesizes new DNA by connecting nucleotides into a chain

joins the new DNA strands together

hydrogen bonds

Faster

untangles the DNA (specifically, DNA gyrase in bacteria)

helicase

primase

polymerase

ligase

Topoisomerase (gyrase in bacteria)

template

5' to 3'

Can only add nucleotides to the 3'OH

replication synthesis

repair synthesis

• by DNA ligase
• joins the 5' phosphate of one DNA molecule to the 3' OH of another to seal fragments together
• seals the backbone between the two Okazaki fragments on the lagging strand

ligase and polymerase

• untangle the DNA strang
• example of a topoisomerase
• present in bacteria cells

5' to 3'

one

one gene-one enzyme

• 1. Mutigenized bread mold spores by emitting them with xrays which caused mutations
• 2. Grew the spores in a complete medium
• 3. Grew them in a minimum medium to try to identify the spores mutated in the ability to create life; if the cell cannot make an amino acid, it is dead
• 4. Added amino acid to the minimum medium; which amino acid, when present, allows them to live?

• The sugar in the backbone: RNA lacks the hydroxyl (-O) on 2'
• DNA is a double helix, RNA is a single strand
• DNA uses ATGC, RNA uses AUGC

RNA is synthesized using DNA as a template

mRNA message is "read" by the ribosome, which directs protein synthesis

group of three nucleotides of mRNA specifies one amino acid

The three-nucleotide combination that specifies a given amino acid

AUG

UAA, UGA, UAG

process by which DNA directs protein synthesis

• DNA is transcribed to make RNA
• RNA is translated to make protein

• quaternary structure means multiple polypeptides
• Ex: hemoglobin is two different genes
• DNA always codes for protein, final product of gene is RNA

transcribed; translated

group of three nucleotides of mRNA that specifies an amino acid

• redundant: many codons will specify the same amino acids
• unambiguous: every time we have GGG, we have glysine no matter what

• Transcribe to RNA= 5' - AAG AUG CCG UGG GUC UCU - 3'
• Start codon = AUG...Met Pro Trp Val Ser

• Initiation
• Elongation
• Termination

• 5' CAP
• 3' Poly A tail
• RNA splicing

• duplication of the entire genome, occurs in S phase
• DNA Polymerase makes DNA from DNA template
• every cell in your body have the same genome but don't express the same genes (ex: Liver cells only express genes for liver cells, skin cells express genes for skin)
• a genome is all the DNA in the cell, a transcriptome is all the genes that are expressed

• only parts of the genome are transcribed
• "transcriptome"
• RNA polymerase makes RNA using DNA template

RNA polymerase is capable of opening the DNA itself

RNA polymerase is capable of beginning transcription without a free 3' end, it can start from scratch

Helicase and primase

"Noncoding" strand

"coding" strand

"coding"/nontemplate strand

3' to 5'

RNAP binds promoter and opens up helix, begins synthesizing RNA

• RNAP continues to synthesize RNA
• reads template strand
• RNA made is almost identical to coding strand (nontemplate)

RNAP reaches specific sequence that prompts the RNAP to fall off DNA

• sequence where RNA Polymerase first attaches to DNA
• 'upstream' sequence that specifies which strand is template (in both prok/euk)
• Eukaryotic promoters have a 'TATA' box motif

indicates which strand is the template strand (not the coding strand)

guild the RNAP to the correct location on the promoter

intervening sequences spliced out of transcript

expressed sequences that are joined during modification

• Helps the export of mRNA from the nucleus
• Protects mRNA from degradation
• Helps ribosomes attach for translation

snRNPs

• recognize sequences of ends on introns
• snips out intron and ligates exons

• Attachment of RNAP to DNA
• Determines which DNA strand is the template

• all the cells in your body have the same genome but don't express the same genes
• Ex: Liver cells only express genes for liver cells, skin cells express genes for skin

• 5' CAP
• 3' Poly A tail
• RNA splicing

in the nucleus; in the cytoplasm

• Replication: duplication of the entire genome, occurs in S phase; DNA Polymerase makes DNA from DNA template
• Transcription: only parts of the genome are transcribed ("transcriptome"); RNA Polymerase makes RNA using DNA template
• Both: enzymes move in 5'-3' direction, adding nucleotides to the 3' end while reading a DNA template

• binds RNA
• carries aminio acid specified by the codon

• mRNA, tRNA: informational
• rRNA, snRNA: catylitic/structural
• SRP RNA: structural
• miRNA, siRNA: regulatory

Translator binds RNA; carries amino acid specified by the codon (aka binds the codon and carries appropriate amino acid as specified by that codon)

complementary to the mRNA codon; when anticodon pairs with codon, it brings with it the appropriate amino acid; attached to the tRNA

third nucleotide in a codon often doesn't matter as much, primary reason why you can get away with 45 tRNAs to bind with 61 codons, third nucleotide doesn't always matter

Elongation

The P site amino acid is attached to the A site amino acid. Then mRNA + tRNAs move one codon over. This continues until a STOP codon is reached. Translation terminations

• targeted to specific locataions
• Signal peptide recognized by SRP (signal-recognition particle) guides ribosome to ER

• Chromosomal rearrangements
• insertions
• deletions
• substitutions
• point mutations

• entire pieces of chromosome misplaced or attached to another chromosome (dramatic)
• Ex: down syndrome-caused by three copies of chromosome 21

• affect one base pair in a gene
• change of a single nucleotide

• Base pair substitution, base pair insertion or deletion
• Result in: silent mutations, missense mutations, nonsense mutations (all on amino acid level)

• arises from base substitution
• no affect on the amino acid sequence because the new nucleotide still codes for the same amino acid, therefore the protein is not affected

• arises from base substitution
• one nucleotide changed that doesn't code for the same amino acid
• can have dramatic consequences (death, cystic fibrosis, sickle cell anemia, etc.)...disease causing

• missense mutation of the amino acid Glu to Val
• Glu is charged/hydrophilic, Val is hydrophobic (carbons in side chain)
• completely changes the shape of the protein (shape is dictated by R groups)
• protein is hemoglobin which carries oxygen around the body shape changed so oxygen is not moved around as efficiently
• odd shape of the hemoglobin makes them stick together to change the shape of the actual cell
• the new shape of the cell doesn't allow it to move through the veins as easily

• arises from base substitution
• premature stop codon being specified (translation stopped, not transcription)
• the protein doesn't get made

insertion or deletion of a nucleotide shifts the frame one basepair, resulting in an 'in frame' stop codon

• Thymine dimer distorts DNA
• Nuclease cuts out damaged DNA
• DNA pol I repair synthesis
• DNA ligase seals the backbone

• changes the shape of the protein if a different amino acid is sensed from a new nucleotide in the sequence
• may go from being hydrophilic to hydrophobic or vice versa

• UV light causes thymine dimers
• when two adjacent T's bind to each other instead of across
• nuclease repairs that bulge

no, they can also be a source of genetic variability in nature and allows animal to adapt to new environments

a premature stop codon is specified so that translation is stopped and the protein doesn't get made

• one promoter for multiple genes
• set of genes coordinately regulated by a single promoter--produces a polycistronic message (one mRNA for more than one gene)

DNA sequence that recruites DNA polymerase

enzyme that breaks down lactose into glucose and galactose

monosaccharides

makes membrane permeable to lactose

• inducer that binds the repressor when lactose is present
• changes the shape and function so that the repressor no longer fits on the DNA and falls off and the RNA polymerase is able to express the transcript
• produced by lacZ (converted by B-gal)
• 1'-4' glycosidic linkage converted to 1'-6' linkage

binding site for the repressor (operator)

repressed by lacI repressor protein bound to operator

• inducer (allolactose) binds repressor and changes its shape
• Repressor falls off the DNA and some low level of gene expression takes place

Glucose inhibits adenylyl cyclase activity resulting in low cAMP levels

• cAMP levels are high and bind with CAP to the promoter (at a specific site) which keeps the RNA polymerase on the DNA
• promotes transcription

• RNA polymerase (bound to promoter)
• cAMP levels high (glucose levels low)

RNA pol falls off all the time, transcription won't be very robust without an activator

cAMP levels are low

• breaks down lactose into glucose and galactose
• rearranges lactose to form allolactose, the inducer

• DNA packaging (access to promoter region is controlled)
• Regulation of transcription initiation
• Post-transcriptional regulation
• Protein degradation

Transcriptional regulation because it saves energy to stop the synthesis of a protein early

• 1. DNA, the double helix
• 2. "beads on a string"-DNA wrapes around histones=nucleosome
• 3. 30-nm fiber
• 4. Looped domains (300-nm fiber)
• 5. Metaphase chromosome

DNA that is always tightly packaged, highly condensed

relatively loosely packaged DNA

Euchromatin

heterochromatin

• Histone tailes can be modified to allow access of RNAP to a gene's promoter
• Tails have a positive charge, DNA has a negative charge creating an electrostatic interaction between these proteins and the DNA
• if we change the charge of the tailes to be negative (acetylation), they repel from the DNA and we now have access to the DNA polymerase
• Add methyl groups to repress expression (makes the DNA pol inaccessible by condensing the DNA (30-nm fiber)

DNA sequences that bind specific transcription factors (like activators) to increase gene expression

• Specific transcription factors bind to the enhancer to stimulate transcription
• General transcription factors bind to proximal control elements to initiate transcription

• Alternative splicing (shuffling exons/introns to create more than one peptide from the same gene)
• Control of mRNA degradation (mRNA stability/RNAi mechanisms)

• polyA tail is chewed up, CAP is removed, nuclease degrades mRNA
• The untranslated region can contain "instability factors" which promote mRNA degradation (when we only need proteins for a short period of time)

• controls half of all gene expression
• Single stranded RNA (called microRNA or small interfering RNA) binds complementary sequences in mRNA
• Results in the repression of translation of mRNA

• Histone tails can be modified to allow access of RNAP to a gene's promoter
• Presence or absence of specific transcription factors control which genes get expressed
• The stability of the transcript and how the transcript is spliced
• Protein (e.g. repressors) can also be degraded if they are no longer needed

loosens association of DNA with histones and allows transcription

some not necessary in others

can block or degrade the translation of specific mRNAs

"instability factors" in the untranslated region promotes mRNA degradation

• Protein is the heritable material
• Protein was transferred from the killed S cells to the R cells
• S cell protein transformed R cells to a virulent strain.

Protein is the heritable material

Chagraff's rules state that in DNA, the percentages of A and T and of G and C are essentially the same, and the fly data are consistent with those rules.

In the Watson-Crick model, each A hydrogen-bonds to a T, so in a DNA double helix, their numbers are equal; the same is true for G and C

some substance from pathogenic cells was transferred to nonpathogenic cells, making them pathogenic

A + G = C + T

An anabolic reaction, creating phosphodiester bonds

phosphodiester

• Ultimately, it's because DNA Poly cannot add to the 5' end so the 5' DNA is never replaced in most cells (a shortened 5' end is produced when primers are removed).
• Bacteria chromosomes are circular, so there's always a free 3' end to which DNA Polymerase can add nucleotides

5'-3' on the newly synthesized DNA

sometimes more than one polypeptide is required to make the final protein (so more than one gene is required for a given protein), also sometimes an RNA is the final product of the gene (ex. tRNA or rRNA)

a three nucleotide unit in mRNA that specifies an amino acid

• There is more than one codon per amino acid (redundant)
• but each codon codes for the same amino acid each time (unambiguous)

a DNA sequence that binds RNA polymerase and initiates transcription

Only a subset of genes are required at any given time

• Both use polymerases that must add nucleotides to the 3' end.
• Replication and Transcription use different enzymes.
• Replication duplicates DNA during S phase of the cell cycle.
• Transcription makes an RNA copy of a gene.

Endergonic

• Pairing at the 3rd position of the codon to the anticodon isn't always strict.
• Inosine in the tRNA can pair with A, C or U.
• Sometimes G's pair with U's.
• The 3rd base wobble is the reason we can get away with 45 tRNAs for 61 codons.

through the action of aminoacyl-tRNA synthetases

a signal peptide in a newly synthesized protein targets the protein to the ER lumen, where it enters the endomembrane system and from there, is targeted to the membrane

• Cells will use glucose first
• Glucose is a monosaccharide

RNA polymerase, cAMP/CAP complex

because RNA polymerase doesn't stay on the DNA very well by itself, so transcription efficiency is relatively low

• two functions:
• a) break down lactose into glucose and galactose
• b) convert lactose into allolactose by rearranging the glycosidic bonds. (B-gal carries out this second function only occasionally)

located in euchromatin

• requires double stranded RNA
• can degrade target mRNAs

tags proteins for degradation

• an example: the proteins that produce skin pigments (melanin) would not be on in the liver cells.
• Proteins that are involved in alcohol detoxification would not be on in skin cells (at high levels)

acetylation opens up DNA, and promotes gene expression

because activators turn on specific genes found only in certain cells

RNAi can degrade mRNAs or prevent translation of mRNAs

tagging proteins with ubiquitin, which leads to protein degradation

Helicase

DNA polymerase III

primase

topoisomerase

• Replication: whole genome (DNA to DNA); DNA pol used; uses nucleotides A,T, C, G;
• Transcription: part of the genome; RNA pol used; uses nucleotides A, U, C, G;

• DNA pol: used to synthesize DNA; does not require helicase; binds to promoters; A, U, C, G
• RNA pol: used to synthesize RNA; requires helicase to unwind DNA; bind to origins of replication; A, T, C, G