Final Exam: New Material Flashcards
1
Q
What are diazotrophs?
A
The only organisms that produce the enzyme, nitrogenase, capable of fixing N2
2
Q
How is ammonium incorporated into biomolecules?
A
A combination of the glutamine synthetase and glutamate synthase reactions results in the formation of an amino acid using ammonium as the nitrogen source.
3
Q
What is the net reaction of incorporating ammonium into biomolecules?
A
aKG + NH4+ + NADPH + ATP –> Glu + NADP+ + ADP + Pi
4
Q
How is nitrogen carried throughout the body?
A
Amino acids serve as the nitrogen carriers within the body. Nitrogen may be incorporated into other biomolecules by incorporating the amino acid.
5
Q
How do amino acids transfer amino groups?
What is a transaminase?
A
Transaminase enzymes swap the carbonyl group of a-keta acids with the amino group of amino acids.
6
Q
What is the nitrogen source for signaling molecules?
A
Many signaling molecules (melatonin, dopamine, etc) are derived from amino acids.
7
Q
Do human beings endogenously produce all 20 amino acids?
A
Animals only have biosynthetic pathways for some nonessential amino acids. Essential amino acids must be consumes through our diet.
8
Q
What does nitric oxide do and what is it derived from?
A
The vasodilator, nitric oxide, is derived from Arginine.
9
Q
How are nitrogenous bases synthesized?
A
Amino acids, THF, and HCO3- are the nitrogen precursors of nitrogenous bases.
10
Q
What is the catabolic fate of purines?
A
Purines are converted to uric acid (urate) and excreted in the urine.
11
Q
What is the catabolic fate of pyrimidines?
A
Pyrimidine catabolism produces intermediates of fatty acid metabolism.
12
Q
What are the products of glucogenic amino acid catabolism?
A
Glucogenic amino acids are catabolized to precursors for gluconeogenesis such as pyruvate and oxaloacetate.
13
Q
What are the products of ketogenic amino acid catabolism?
A
Ketogenic amino acids are catabolized to acetyl-CoA for use in ketogenesis or fatty acid synthesis.
14
Q
How do ammonotelic organisms (crustaceans and some fish) excrete excess nitrogen?
A
Ammonotelic organisms excrete nitrogen as ammonia (NH3), requires a lot of water but uses the least energy.
15
Q
How do Ureotelic organisms (mammals and some reptiles) excrete excess nitrogen?
A
Ureotelic organisms excrete nitrogen as urea, requires some energy investment but only a little water.
16
Q
How is urea made?
What enzyme catalyzes this reaction?
A
Amino acids transfer their amino groups to a-ketoglutarate using a transaminase reaction to form Glu and Asp which donate their Nitrogen groups to form urea.
17
Q
How do Uricotelic organisms (birds and reptiles) excrete excess nitrogen?
A
Uricotelic organisms excrete nitrogen as uric acid, requires very little water but more energy to make.
18
Q
Why is using less water to excrete nitrogen an advantage to birds?
A
Means less weight
19
Q
Why is using less water to excrete nitrogen an advantage to animals in the desert?
A
Less water loss
20
Q
What is the Cori cycle?
A
Under anaerobic conditions, muscles produce lactate which is exported to the liver for gluconeogenesis. The liver then exports the glucose back to the muscles.
21
Q
Define hormones.
A
Hormones are substances produced by one tissue that affect the function of other tissues throughout the body.
22
Q
What does insulin do and where is it released from?
A
Insulin is released by the B-cells of the pancreas in response to high blood glucose. It signals fuel abundance promoting fuel storage while limiting the release of stored fuels.
23
Q
How does insulin regulate fuel metabolism in muscle tissue?
A
-promotes glucose transport into cells
-stimulates glycogen synthesis
-suppresses glycogen breakdown
24
Q
How does insulin regulate fuel metabolism in adipose tissue?
How does it affect ACC?
A
-Activates extracellular lipoprotein lipase
-Increases level of acetly-CoA carboxylase
-Stimulates triacylglycerol synthesis
-Suppresses lipolysis
25
How does insulin regulate fuel metabolism in the liver?
| Promotes fuel storage and inhibits new fuel breakdown
-Promotes glycogen synthesis
-Promotes triacylglycerol synthesis
-Suppresses gluconeogenesis
26
How does insulin stimulate glucose import?
Insulin receptor initiates a signaling cascade with various intracellular effects including the fusion of vesicles containing the GLUT4 transporter, with the plasma membrane.
27
How does insulin regulate fuel storage?
Insulin signaling activates phosphatases that dephosphorylate both glycogen synthase (activating it) and glycogen phosphorylase (inhibiting it).
28
What is glucagon?
Glucagon is a small peptide hormone released by the a-cells of the pancreas in response to low blood glucose and opposes the effects of insulin.
29
How do glucagon and epinephrine regulate fuel metabolism on liver cells?
| What do they promote?
Epinephrine and glucagon have similar effects on liver cells (they promote gluconeogenesis and glycogen breakdown).
30
How do glucagon and epinephrine regulate fuel metabolism on muscle cells?
Muscle cells do not have glucagon receptors or the enzymes for gluconeogenesis. Epinephrine promotes glycolysis and glycogen breakdown.
31
What is diabetes mellitus?
A disorder of fuel metabolism characterized by high blood glucose.
32
What is Type 1 diabetes? How is it treated?
Type 1: Autoimmune disease that kills pancreatic B-cells. Treated with insulin injections.
33
What is Type II diabetes? How is it treated?
Type 2: Insulin resistance. Treated with lifestyle changes, drugs that lower blood glucose, or bariatric surgery.
34
What are the long term affects of diabetes?
Hyperglycemia leads to long term effects such as cataracts, kidney failure, and cardiovascular damage.
35
Why is DNA replicated in a semiconservative manner?
In a new DNA helix, 1 strand is old and 1 strand is new.
36
How is DNA replication intiated?
Proteins associated with a specific DNA sequence called an origin of replication open up an initial replication bubble.
37
What does the DNA helicase do?
| What kind of energy does it use?
Helicase uses ATP to unwind the DNA helix.
38
What does the Single-strand binding protein (SSB) do?
SSB binds the single-stranded regions to prevent reannealing or nuclease degradation of the DNA.
39
What does the DNA polymerase do?
DNA polymerase adds deoxynucleotides to the free 3' OH of an existing strand.
40
Draw the DNA polymerase mechanism.
41
What is the main prokaryotic polymerase and how many polymerases does E. coli have?
E. Coli has 5 DNA polymerases. DNA pol III is the main polymerase used for replication.
42
What is the main eukaryotic polymerase(s) and how many polymerases do humans have?
Humans have at least 14 DNA polymerases. DNA pol S does lagging strand synthesis and DNA pol E does leading strand synthesis.
43
What happens if DNA polymerase adds an incorrect base?
| What direction is it fixed?
Most DNA polymerases have a proofreading function: the polymerase slows down due to the bulge caused by the mispairing and a 3' to 5' exonuclease removes the misincorporated nucleotide.
44
What direction does DNA polymerase run?
5' to 3'
45
Why can't the DNA polymerase extended DNA in the opposite direction (3' to 5')?
If you make a mistake an have to remove a group at the 5' end, there is only 1 phosphate group left which doesn't leave enough energy to continue polymerization.
46
What does the primase do?
Introduces a short RNA primer that is complementary to the template strand
47
What happens to the RNA primers during replicatoin?
A 5' to 3' exonuclease (RNase H or DNA pol I) removes the RNA primer from each Okazaki fragment.
48
What does the DNA ligase do?
DNA ligase forms a phosphodiester bond to join neighboring fragments.
49
What are telomeres?
The ends of chromosomes adopt special structures called telomeres where a repeating sequence of DNA folds into a loop and binds protective proteins.
50
What is the end-shortening problem? What is the solution?
Linear DNA gets shorter after a round of replication as the 5' RNA primer cannot be replaced. Telomerase adds 5' DNA repeats to the 3' ends of human DNA.
51
What is telomerase?
Telomerase is a reverse transcriptase that carries an RNA template that codes for a new repeat while base-pairing with the previous repeat. After a round of replication, the shortened chromosome is missing repetitive DNA rather than essential genetic information.
52
How is DNA packaged?
| What is a nucleosome?
Linear DNA (string) is negatively supercoiled by wrapping the double helix around a core of 8 histones to form a nucleosome (beads).
53
What is euchromatin?
Lightly packed DNA that is heavily transcribed.
54
What is heterochromatin?
Tightly packed DNA that is inaccessible for DNA transcription.
55
What are ways that DNA can be damaged?
DNA is subject to oxidative damage, spontaneous hydrolysis, and methylation.
56
What is a Guanine --> 8-oxoguanine form of DNA damage?
Caused by reactive oxygen species such as peroxide and results in a G to T transversion.
57
What is a transversion point mutation?
A transversion is a mutation leading to a switch between purine and pyrimidine.
58
What is a transition point mutation?
A transition is a purine to purine or pyrimidine to pyrimidine mutation.
59
How does spontaneous hydrolysis cause an abasic site?
The result of water attaching to the anomeric carbon and clipping off the nitrogenous base.
60
What is a deamination form of DNA damage?
Cytosine undergoes a deamination reaction with water and turns into a Uracil.
61
What is Base Excision Repair?
| What is a glycosylase?
Cells have specific glycosylases that recognize common errors and remove the incorrect base.
62
Describe the process of Base Excision Repair.
1. DNA glycosylase removes the incorrect base creating an abasic site.
2. An Endonuclease breaks the backbone.
3. DNA polymerase and DNA ligase fix the strand.
63
Why is sunshine mutagenic?
UV light promotes pyrimidine dimers which disturbs the DNA shape.
64
How do environmental chemicals such as benzopyrene cause DNA damage?
Benzopyrene is oxidized in the liver and causes a G-->T transversion which distorts the helix.
65
What is nucleotide excision repair?
Errors that significantly distort the helix are repaired by removing a segment of DNA surrounding the distortion.
66
What is the process of nucleotide excision repair?
1. A helicase separates the damaged strands
2. An endonuclease removes the region around the damaged nucleotide
3. DNA polymerase and DNA ligase repairs the strand.
67
How is DNA methylation repaired?
Suicide proteins called alkytransferases, transfer the methyl group to a cysteine (permanently inactivating the enzyme).
68
How are misincorporated bases repaired?
After replication, the mismatch repair system scans the newly synthesized strand and excises any segments with misincorporated nucleotides.
69
What happens in nonhomologous end-joining?
| What proteins are recruited?
A double strand break is recognized by Ku proteins which recruit a nuclease that trims some nucleotides, a DNA polymerase that adds some nucleotides, and a ligase that connects the backbone.
70
What can induce a double strand-break?
Radiation or free radicals
71
What happens in homology-directed repair?
If a second chromosome is present it will be used as a template to repair the broken chromosome.
72
How can we cut DNA at specific locations?
| What is special about the sequence that is cut?
Restriction Enzymes are isolated from bacteria and cut double-stranded DNA within a defined recognition sequence that is usually palindromic.
73
What is the overhang used for after cutting DNA?
Complementary sticky ends allow reannealing and ligation of DNA fragments.
74
Define a plasmid.
Plasmids are small circular DNAs that replicate independently in bacteria.
75
What are the characteristics of plasmids?
1. Have an origin of replication
2. Have an antibiotic resistance gene to select for bacteria carrying the plasmid
3. Include translational and transcriptional control sequences
4. Have a polylinker region
76
What is a polylinker region in a plasmid?
A set of unique restriction enzyme sites that allow directional insertion of DNA fragments
77
What are the steps for producing recombinant DNA?
1. Cut the gene and the vector using restriction enzymes that generate the same sticky ends
2. Use gel electrophoresis to separate unwanted fragments
3. Use complementary sticky ends to anneal the gene inside a vector
78
How does cloning produce recombinant DNA?
Cloning involves the recombination of DNA. A polypeptide of interest can be expressed by inserting a gene into a vector that may be replicated by the host organism.
79
What is polymerase chain reaction (PCR)?
PCR is a technique that amplifies any DNA sequence provided some information is known on either side of the target DNA to be amplified.
80
What does a PCR need to work?
Need template DNA, dNTPs, primers that flank the sequence, Mg2+, and a thermostable polymerase
81
What are the 3 steps of PCR?
1. ~95*C --> DNA is denatured
2. ~60*C --> Primers anneal
3. ~72*C --> DNA polymerase extends the primers.
82
What is the end result of PCR?
DNA is effectively doubled each cycle.
83
What are the steps for Illumina sequencing?
84
How do next-generation sequencing methods work?
Genomic DNA is fragmented into manageable pieces and known sequences are attached to the ends of each fragment.
85
What is added each cycle of Illumina sequencing?
Each cycle, dATP, dCTP, dGTP, and dTTP are added with each deoxynucleotide connected to a different fluorescent group.
86
What does the fluorescent color of each deoxynucleotide indicate?
At each spot, the fluorescent color indicated which base was added to the growing strand opposite the unknown fragment and a computer records the identity of the nucleotide.
87
How are the fluorescent groups removed after the nucleotide is added?
Fluorescent groups are chemically cleaved and washed away leaving a free 3' OH group before beginning the next cycle.
88
How are entire genomes sequenced?
Overlapping sequences from different fragments may be assembled to sequence entire genomes.
89
How do bacteria recognize an invading virus?
Bacterial chromosomes have segments of viral DNA within a set of clustered regularly interspersed short palindromic repeats (CRISPRs)
90
How do bacteria know which sequences to cut from viruses?
Bacteria transcribe the viral segments and the Cas9 protein uses the resulting guide RNAs to locate and destroy and complementary DNA sequences that might be present in an invading virus.
91
How can we alter the DNA in a cell using Cas 9 and nonhomologous end-joining? Why would you use this technique?
Targeted genes may be inactivated by Cas9 as nonhomologous end-joining is inherently mutagenic resulting in a nonfunctional protein. You can observe the cell when it can't make the protein in order to study the function of the protein.
92
How can we alter the DNA in a cell using Cas 9 and homology-directed repair?
Genes may be edited by homology-directed repair after Cas9 cleavage, if replacement DNA is introduced. The cell will repair the cut DNA based on the replacement DNA sequence (that is modified to produce a desired protein sequence).
93
What is a gene?
A gene is a segment of DNA that codes for a protein via mRNA or another functional RNA.
94
What does RNAP1 do?
Produces ribosomal RNA (rRNA) that includes structural and catalytic components of the ribosome.
95
What does RNAPII do?
Produces messenger RNA (mRNA) which codes for polypeptides.
96
What does RNAPIII do?
Produces rRNA and transfer RNA (tRNA) which delivers amino acids during translation.
97
What does an RNA polymerase do?
RNA polymerase synthesizes new RNA strands from a DNA template.
98
What is acetylation of histones?
| What residue it added to?
Acetylation of histone Lys residues typically loosens the DNA-histone interaction and promotes transcription.
99
What other covalent modifications of histones other than acetylation happen?
Other histone modifications, such as phosphorylation, methylation, and ubiquitination have varying effects on chromatin structure and transcription that depend on the specific residue being modified.
100
How does the eukaryotic RNA polymerase know where to start?
Eukaryotes have a variety of upstream and downstream elements that vary from gene to gene. General TFs bind some combination of these elements and recruit RNAPII to the start site.
101
What distal regulatory elements affect transcription?
Enhancers and silencers are DNA sequences that help regulate transcription.
102
What are repressors?
Proteins that decrease the activity of RNAP.
103
What are activators?
Proteins that increase the ability of RNAP to transcribe a gene.
104
How are activators that may be thousands of base pairs away connected to the transcription complex?
Protein complex called mediator.
105
What general transcription factors are always required to initiate transcription?
Eukaryotic transcription initiation requires 6 highly conserved general transcription factors that help RNAPII locate the transcription start site and open up the transcription bubble.
106
What does TFIID do?
Includes the TATA-binding protein and associated factors that help recruit TFIIIB.
107
What does TFIIA do?
Assists TATA-binding protein.
108
What does TFIIB do?
Helps recruit TFIIF-RNAPII to the transcription site.
109
What does TFIIF do?
| How does it stabalize the transcripiton bubble?
Increases RNAPII's affinity for the start site, elongate the mRNA product, and bind the nontemplate strand to stabilize the transcription bubble.
110
What does TFIIE do?
| What does TFIIH do?
Recruits TFIIH which acts as a helicase to open up the transcription bubble.
111
How is the RNA transcript elongated?
| Transcription not translation.
Phosphorylation by ATP of the C-terminal domain allows RNAPII to dissociate from mediator and some of the general transcription factors so that it can make the full-length transcript.
112
What is intrinsic termination in E. coli?
In E. coli, transcription may be intrinsically terminated when RNAP transcribes a region that pulls apart the RNA-DNA hybrid helix when the RNA base pairs with itself to form a hairpin structure.
113
What is Rho termination in E. coli?
Transcription may be terminated in E. coli by a Rho protein factor that specifically binds a C-rich region on the transcript and bumps RNAP off the template.
114
What is a bacterial operon?
Operons consist of several genes under the control of one promoter that are transcribed as one mRNA.
115
How can eukaryotes regulate the expression of some groups of related genes?
Related genes may be grouped together under the control of the same regulatory elements but are transcribed separately.
116
How is the 5' end of eukaryotic mRNAs protected from exonucleases?
| Via what linkage?
Eukaryotic mRNAs are capped at the 5' end by linking a guanosine residue to the emerging mRNA via a 5'-5' triphosphate linkage.
117
How is the 3' end of the eukaryotic mRNAs protected from exonucleases?
Eukaryotic mRNAs are polyadenylated at the 3' end. polyA binding proteins protect the mRNA from degradation and help control the mRNA lifetime.
118
What are introns and exons?
Eukarytoic mRNA is transcribed with exons (expressed regions), that code for the polypeptide sequence, separated by introns that are eventually spliced out.
119
How does the cell know where to splice the exons?
Splicing occurs at conserved sequences at the exon-intron junction
120
How does the cell remove introns?
Splicing occurs through two transesterification reactions catalyzed by the splicesome or catalyzed by the RNA itself
121
What are the 2 steps for removing an intron?
1. 2' OH group of the branch point adenosine on the intron attacks the Phosphate at the 5' at the end of the intron. The frees the 3' end of the first exon
2. The free 3' OH group of the first exon attacks the 5' phosphate of the second exon. This reaction forms a phosphodiester bond to unite the 2 exons.
122
What is the advantage of splicing?
Most human structural genes undergo alternative splicing where different processing of the transcript produces variant mature mRNAs.
123
How can RNA act as an enzyme?
RNA has multiple functional groups and adopts unique 3D conformations just like a protein enzyme (splicesome).
124
What is a frameshift mutation?
Insertions or deletions of a number of nucleotides that isn't evenly divisible by 3.
125
How are codons translated to polypeptides?
Codons are recognized by the 3 base anticodon in tRNA. mRNA is read by the ribosome in the 5'-3' direction and protein synthesis proceeds from N-term to C-term.
126
What is a point mutation?
A point mutation involves a single base substitution
127
What is a nonsense mutation?
A point mutation that in which the new codon codes for stop.
128
What is a silent mutation?
A point mutation in which the new codon codes for the same amino acid.
129
What is a missense muation?
| What are the subtypes?
A point mutation in which the new codon codes for a different amino acid.
-conservative: the new codon codes for a similar amino acid
-nonconservative: the new codon codes for a dissimilar amino acid
130
What is a tRNA?
tRNAs serve as adapters between mRNA and amino acids.
131
What is the structure of a tRNA?
The tRNA anticodon base pairs with the codon to read the mRNA while the acceptor stem is linked to the appropriate amino acid
## Footnote
The anticodon reads 5' to 3', while the mRNA reads 3' to 5'
132
How can a single anticodon read multiple codons?
The 5' anticodon position experiences some flexibility (wobble) in its bonding with the 3' condon position.
| Changes in the 3' codon base usually code for the same amino acid.
133
How is an amino acid attached to the correct tRNA?
Aminoacly-tRNA synthetases (aaRS) are enzymes that add the correct amino acid to a given tRNA.
## Footnote
Each aaRS recognizes a specific amino acid along with all associated tRNAs for that amino acid
134
What is the ribosome?
| What kind of bonds does it form?
A protein-RNA complex that catalyzes the formation of peptide bonds.
135
# The ribsome has 3 sites between the large and small subunits.
Describe the 3 sites of the ribosome.
- The aminoacyl (A) site binds the incoming aminoacyl-tRNA
- The peptidly (P) site binds the tRNA with the growing polypeptide chain
- The exit (E) site transiently binds the exiting deacylated tRNA
136
How is the start codon located for translation in prokaryotes?
RNA within the 30S subunit binds the Shine-Dalgarno sequence upstream of the start codon on the mRNA.
137
# In prokaryotic translation:
What does Intiation Factor 2 (IF-2) do?
| What is blocked?
Brings the charged intiator tRNA to the start codon in the P-site. The A-site is blocked by IF-1.
138
# In prokaryotic translation:
How is the polypeptide elongated?
EF-Tu delivers a new aminoacyl-tRNA to the A site.
## Footnote
If the correct codon-anticodon match is made, the ribosome will shift its conformation to ineract with the first 2 base pairs and hydrolysis of GTP. EF-Tu GDP departs
139
Draw the mechanism of polypeptide elongation.
140
# After EF-Tu brings a new amino aicd,
How is the polypeptide elongated?
EF-G bumps the peptidyl-tRNA from the A site to the Psite and the deacylated tRNA from the P site to the E site.
| 1 bump moves the mRNA by 3 nucleotides -introduces a new codon in site A
141
How is translation terminated?
| What is needed to prepare for another round of peptide synthesis?
Ribosome recycling factor (RRF), EF-G, and IF-3 remove the deacylated tRNA and separate the ribosomal subunits in preparation for another round of peptide synthesis.
142
What factors recognize the stop codon?
Releasing Factor 1 (RF-1) and RF-2 causes the ribosome to transfer the peptidyl group to water to release the polypeptide chain.
143
What is the signal recognition particle (SRP)?
| What happens to proteins destined for the plasma membrane?
The signal recognition particle (SRP) recognizes amio acid seqeunces being translated with a positively charged residue followed by several hydrophobic residues and stops translation and brings it to the ER for eventual secretion or direct insertion to membranes.
144
Describe the intiation of polypeptide from mRNA in detail.
Intiation requires an initiation factor (IF2) bound to an initial tRNA with fMet. The initial tRNA anticodon locates the start codon of the mRNA and binds the small ribosomal subunit which is positioned by the Shine-Dalgarno sequence in bacteria. IF1 binds in the A-site to prevent premature tRNA binding in this location. Binding of the large ribosome subunit promotes dissociation of IF1 and hydrolysis of the GTP associated with IF2.
145
Describe the elongation of a polypeptide from mRNA in detail.
EF-Tu delivers the next charged tRNA to the ribosome A site. If the codon-anticodon pairing is correct, then EF-Tu hydrolyzes GTP and leaves the tRNA behind in the A-site. EF-G then pushes the A-site tRNA into the P-site and bumps the now empty tRNA from the P-site into the E-site. EF-G hydrolyzes GTP and dissociates.
146
Describe the termination of a polypeptide from mRNA in detail.
Once the stop codon is in the A-site, a protein release factor binds and promotes the transfer of the peptidyl group from the P-site to water. The peptide then exits the ribosome.RF3 hydrolyzes GTP to remove RF1/2. EF-G and a ribosome recycling factor (RRF) then bind to remove the deacylated tRNA from the P-site and IF-3 helps separate the ribosomal subunits.
147
How do bacteria expressing restriction endonucleasers protect their own DNA from digestion?
Bacterial DNA is methylated at restriction sites.
148
Describe self-splicing introns.
RNA of the intron folds over into a structure that can bind a free Guanosine then orient its hydroxyl to attack the phosphodiester bond between the exon and intron.
