Semester 1: Midterm 2 Flashcards
What is he basic unit of a lipid?
A fatty acid: carboxylic acid with hydrocarbon chains ranging from C4-C24
What is a saturated fatty acid?
A fatty acid with no double bonds
The double bond of unsaturated fatty acids is usually in … confirmation
cis (trans can be found in bacteria)
What determines the boiling point of a fatty acid?
The more double bonds -> more soluble -> higher surface area -> higher boiling point
What are the essential fatty acids?
Arachidonic acid, linoleic acid, linolenic (two last used to make the first)
What is the composition of TAGs?
Three fatty acids in ester linkage with a single glycerol
What are the basic unit of cholesterol?
Sterols (structural lipids present in the mem. of most eukaryotic cells)
- Have 4 fused rings and a hydroxyl group
What are the reduced derivatives of cholesterol?
Bile acids
Primary bile acid
- 24 carbons
- 2-3 hydroxyl groups
- Side chain that terminates in a carboxyl group
Secondary bile acid
Produced from primary bile acids by removing on of the hydroxyl groups
When does ARA (arachidonic acid) become essential in the diet?
If linoleic acid is deficient in the diet
What are eicosanoids?
The collective name for prostaglandind and the related compounds thromboxanes and leukotrienes
What are nucleotides composed of?
1) A nitrogenous base (adenine, guanine, cytosine, thymine, uracil)
2) A pentose monosaccharide
3) 1-3 phosphate gorups
What is a nucleoside composed of?
Base + pentose sugar
The backbone of the nucleic acid is made up from…?
Covalent phosphodiester bond between the nucleotides, going from3’ to 5’ end
How is adenine and thymine connected in DNA?
By two hydrogen bonds (+hydrophobic interactions)
How is cytosine and guanine connected in DNA?
By three hydrogen bonds (+hydrophobic interactions)
Structural forms of the double helix: A, B and Z forms
- B-form: Right handed helix with 10 residues per 360 degree turn of the helix (normal form)
- A-form: Dehydrated form of beta-form (11 per turn)
- Z-form: Left handed (12 per turn)
Differences between RNA and DNA
- RNA is smaller
- RNA contains ribose instead of deoxyribose
- RNA contains uracil instead of thymine
- RNA exists as a single strand
What are exonucleases?
Enzymes that work by cleaving nucleotides one at a time from the end of a polynucleotide chain
What are endonucleases?
Enzymes that cleave the phosphodiester bond within a polynucleotide chain
What are the functions of DNA polymerase I?
(Prokaryotes) Gap filling, following DNA replication, repair and recombination
- 3’-5’ exonuclease (proofreading)
- 5’-3’ exonuclease: Removes RNA primer when Pol III is in close proximity to it
- 5’-3’ polymerase activity
What are the functions of DNA polymerase II?
(Prokaryotes) DNA proofreading and repair
Requires duplex DNA template and primer
- 3’-5’ exonuclease
What are the functions of DNA polymerase III?
(Prokaryotes) Functions as the replication fork
- Catalyzes the highest rate of chain elongation in 5’-3’ direction
- 3’-5’ exonuclease: Proofreading
What is the function of DNA ligase?
Catalyzes the formation of a phosphodiester bond between a 3’hydroxyl at the end of one DNA strand and a 5’phosphate at the end of another strand
What proteins makes up the prepriming complex?
- DnaA protein: Binds to specific nucleotide sequence at origin of replication giving AT-rich regions in the origin to melt
- DNA helicase: Forcing dsDNA apart (unwinding double helix)
- SSB proteins (single-stranded DNA-binding): Cooperative binding to ssDNA - keeps strands apart
Function of topoisomerase
Unlinks parental strands
What is leading and lagging strand?
- Leading: Strand being copied in 5’-3’ direction (forward), continuously
- Lagging: Retro-grade, DNA synthesized in short segments (Okazaki-fragments)
Function of Polymerase alpha?
(Eukaryotes)
- Contains primase -> synthesize a short RNA primer
- Initiates DNA synthesis on leading strand
Function of Polymerase beta?
(Eukaryotes)
- Repair (gap filling)
Function of polymerase gamma?
(Eukaryotes)
- Replicates mitochondrial DNA
Function of polymerase delta?
(Eukaryotes)
- Elongation of Okazaki fragments on lagging strand
Function of polymerase epsilon?
(Eukaryotes)
- Elongation of leading strand and complete DNA synthesis on this strand
Why do we have replication bubbles in eukaryotes?
1) Replication is bidirectional
2) Replication proceeds from multiple origins in each chromosome
Nucleosome structure
Histones and DNA:
- Two H2A, H2B, H3 and H4 form the structural core of the individual nucleosome “beads” (histone octamer)
- A segment of double helix DNA wound around core
DNA dependent RNA polymerase complex and each subunits function
Consisting of:
- 2 identical alpha subunits (binds regulatory sequences)
- 2 large beta (forms phosphodiester bonds) and beta’ (binds DNA template) subunits
- Omega subunit
= often termed E -> associates with sigma to form holoenzyme (sigma factor allows RNA pol to recognize initiation site)
What are the promoter regions recognized by RNA pol sigma factor?
- Pribnow box
- 35 sequence
Termination of transcription in prokaryotes
1) Rho-dependent: Rho factor migrates along behind RNA pol in 5’-3’ direction until termination site is reached -> dissociation
2) Rho-independent: Requires that RNA has two structures, stable hairpin turn (with C and G rich sequence) and string of U’s which when binding to A’s of DNA template becomes weak -> separation
Inhibitors of transcription
- Rifampicin (binds beta subunit of bacterial RNA polymerase, blocks promotor clearance (elongation))
- Actinomycin D (intercalates between successive G=C base pairs in duplex DNA)
- Alpha amantin (inhibitor of RNA pol II)
Promotor regions in eukaryotic transcription
- TATA/Hogness box (25n upstream)
- CAAT box (70-80n upstream)
RNA pol I function
(eukaryotes)
Synthesizes the precursor of the large ribosomal RNAs (28s, 18s and 5.8s) in the nucleolus
RNA pol II function + inhibitor
(Eukaryotes)
Synthesize the precursors of mRNA’s
Inhibited by alpha-amantin
RNA pol III function
(Eukaryotes)
Produces the small RNAs, including tRNA
How is the primary transcript of rRNA modified?
Cleaved by ribonucleases to yield intermediate sized pieces of rRNA
- Methylation of pre-rRNA (45S)
- Cleavage to form mature rRNA: 18S, 5.8S, 28S
How is the primary transcript of tRNA modified?
An intron must be removed from the anticodon loop, and sequences at both 5’ end and 3’ end of molecule must be trimmed
How is the primary transcript of mRNA (hnRNA) modified?
1) 5’capping: 7-methyl-guanosine attached backwards to 5’ terminal end of mRNA
2) Poly-A tail: Addition of 40-200 adenine nucleotides to 3’end
3) Removal of introns
Mechanism of splicing (removal of introns from mRNA primary transcript)
1) Primary transcript combines with snRNPs to form folded complex called a spliceosome
2) The 2’OH of an adenosine residue in the intron attacks and forms phosphodiester bond with the phosphate at the 5’ end of intron 1
3) Newly freed 3’OH of upstream exon 1 forms phosphodiester bond with 5’ end of downstream exon 2
4) Intron released
What are the subunits of eukaryotic and prokaryotic ribosomes?
Prokaryotic: 50S + 30S = 70S
Eukaryotic: 60S + 40S = 80S
Binding sites on ribosomes for tRNA
- A site: Binds incoming aminoacyl-tRNA during translation
- P site: Occupied by peptidyl-tRNA -> carries chain of AA that has already been synthesized
- E site: Occupied by the empty tRNA as it is about to exit the ribosome
Components needed for initiation of protein synthesis
1) The 2 ribosomal subunits
2) The mRNA to be translated
3) The aminoacyl-tRNA specified by the first codon in the message
4) GTP
5) Initiation factors facilitating the assembly of this initiation complex
How does the ribosome recognize the sequence initiating translation?
- Shine-Dalgarno sequence: Purine rich sequence, 6-10 bases upstream of AUG codon (prokaryotes)
- Eukaryotes: 40S subunit binds cap structure at 5’ end of mRNA
Role of eukaryotic initiation factors: eIF2
GTP-binding protein interacts with eIF3 and members of eIF4. Binds tRNA
Role of eukaryotic initiation factors: eIF3
Binds to ribosomal subunit-mRNA complex
Preventing large ribosomal subunit from binding small subunit before it’s ready to commence elongation
Role of eukaryotic initiation factors: eIF4
Unwinds tRNA at 5’end
Elongation factors in prokaryotes:
- EF-T4 : Mediates entry of aminoacyl-tRNA to a free site of ribosome
- EF-T5 : Guanine nucleotide exchange factor for EF-T4. Catalyzing the release of GDP from EF-T4
- EF-6 : Catalyzes translocation of tRNA and mRNA down the ribosome at the end of each round of polypeptide elongation
Elongation factors in eukaryotes:
- eEF-1: alpha- entry of aminoacyl-tRNA to free site of ribosome. beta4- Guanine nucleotide exchange factor for alpha, catalyzing release of GDP
- eEF-2: Catalyzing translocation of the tRNA and mRNA down the ribosome at the end of each round of polypeptide elongation
What is a signal sequence?
A short (3-60AA long) peptide chain that directs the transport of a protein
What is co-translational translocation?
The process of exporting proteins from the cell through the rER membrane
The process of co-translational translocation
1) N-terminus of signal sequence being recognized by SRP (signal recognition particle)
2) Protein-ribosome complex transferred to an SRP receptor on ER (2 binding sites: one for signal sequence and one for A-site in ribosomes)
3) Protein inserted into translocon (Sec61 channel- mem. bound protein)
4) Protein covered by a chaperone protein - folded and modified
What is KDEL?
Retention signal sequence for ER resident proteins
What are the role of chaperones in the ER?
Helps in folding, refolding of denatured proteins, preventing misfolded proteins from leaving the ER
What is the role of SNARE proteins?
Mediate fusion of vesicles with membranes
v-snare on vesicle, t-snare on membrane
What is the role of coat proteins? Types?
Vesicle producing proteins
- COPI: Golgi -> ER (retrograde transport)
- COPII: ER -> Golgi (anterograde transport)
- Clathrin: Golgi -> lysosomes
What type of bond keeps nucleotides together?
3’-5’ phosphodiester bonds
What is the correct base pairing for mRNA from this noncoding/antisense (template) strand: CGTCATGTA
GCAGUACAU
How is RNA polymerase different from DNA polymerase?
- Only works on one strand
- Has its own helicase activity
- Does not need priming (primase)
- Does not proofread product (RNA)
- RNA doesn’t stay bound following synthesis
All DNA polymerase a catalyze elongation of the primer strand in …… direction, copying the template strand in a….. direction
5’ to 3’, 3’ to 5’
Start codon for translation
AUG (codes for met)
Stop codon for translation
UAA, UAG, UGA
What causes a frame shift mutation?
Either an insertion or a deletion of a nucleotide
Ames test
- Culture of his- Salmonella
- Medium lacking histidine
Amino acid activation for tRNA
AA+tRNA+ATP –> Aminoacyl-tRNA+AMP+2Pi
Single base changes (point mutations)
- Transitions: Pyrimidine/purine changed to different pyrimidine/purine
- Transversions: Pyrimidine to purine or the other way around
What is a missense effect?
Occurs when a different AA is incorporated at the corresponding site in the protein molecule
What is a nonsense codon?
Can occur after a point mutation, will lead to the premature termination of translation
Cyclin D
- Appears in late G1 phase and allows progression to S phase
- Activates CDK4 and 6 (together with cyclin D forming an active serine-threonine protein kinase)
Function of Rb protein (retinoblastoma protein)
Binds and inactivates E2F
Cyclin E, A and CDK2
Initiation of DNA synthesis in early S phase
Cyclin B and CDK1
Transition from G2 to M phase
DNA double strand breaks can be repaired by..
- Homologous recombination (HR) –> S, G2 and M phases
- Nonhomologous end-joining (NHEJ) –> G0/G1 phase
Tumor suppressor p53 function
Delays transit through cycle (checkpoints G1 and 2)
p21 function
CDK-cyclin inhibitor
