Metabolism 2 (part 2) Flashcards
If the carbon skeleton of an amino acid, after losing its amino group is degraded into acetyl CoA, oxaloacetate, and/or ketone bodies, what is it called?
Ketogenic
ketone bodies for ketogenesis
What are the 2 ketogenic amino acids?
LL
Lysine and Leucine
How many essential amino acids are there?
What does it mean to be essential?
10 out of 20
we can’t synthesize essentials
How many non-essential AA’s are there?
Is arginine essential?
10
Arginine sometimes considered essential, because we don’t synthesize enough
Is tyrosine an essential AA?
No, it’s considered non-essential because we make it from phenylalanine.
However, phenylalanine IS essential
What is tyrosine for an individual incapable of metabolizing phenylalanine?
Conditionally essential
Name the non-essential AA’s
PVT TIM HLL
What is the term to define an amino acid losing its amino group, and the leftover carbon skeleton entering GNG pathway?
Glutogenic
What is gastric acid and from where is it secreted?
What is its function?
HCl
secreted from parietal cells in the stomach
denatures proteins (begins process) and releases intrinsic factor so B12 can be absorbed
What type of hormone is Cholecystokinin?
What is its function?
Peptide hormone
digests Fat AND Protein, stimulates gall bladder, slows gastric emptying, satiety signal
What zymogen stimulates the release of a basic solution from the pancreas?
How long is the zymogen and active form?
Secretin
121AA cut to 27 AA hormonal active form
What enzyme cleaves trypsinogen to become trypsin?
Enteropeptidase
How is trypsin activated?
What does it do?
Trypsinogen > (Enteropeptidase) > Trypsin
Trypsin cleaves chymotrypsinogen > chymotrypsin
What are the 3 main zymogens in protein digestion?
Pepsinogen
Chymotrypsinogen
Trypsinogen
What zymogen is released by chief cells in the stomach in response to HCl?
Pepsinogen
HCl activates Pepsin
Where is chymotrypsinogen made and what does its active form break down?
Pancreas
Chymotrypsin breaks down aromatic AA residues
What activates trypsinogen (trypsin)?
What does trypsin activate?
Enteropeptidase
Chymotrypsin (from chymotrypsinogen)
Elastase (from proelastase)
Carboxypeptidase (from procarboxypeptidase)
Lipase (from prolipase)
What do trypsin, chymotrypsin, and elastase preferentially cleave?
trypsin - arg/lys residues
chymotrypsin - aromatic AA residues
elastase - hydrophobic AA’s
After proteins are sufficiently broken down in the lumen how do they get into the enterocyte?
Secondary Active Transport (specific carrier-mediated)
What size are the AA chains that enter the enterocyte from the lumen?
Free AA, di, and tri-peptides
1-3
What catalyzes the cleavage of AA’s from the N-terminus of oligopeptides entering the enterocyte?
Aminopeptidase
Once 1-3 AA chains are in the enterocyte, what occurs before they are released into the blood?
How are they released into the blood?
Peptidases continue to break into single AA’s
Facilitated transport
What determines the lifespan of a protein?
The AA at the N-terminus
What determines the shelf life of a protein?
PEST sequences (Proline, Glutamate, Serine, Threonine)
AA sequences that serve as marker for death - more protein has shorter its life
What death marker attaches to Lysine?
Ubiquitin?
What structure does the ubiquitin-attached protein enter for degradation?
What survives?
26S Proteasome
Ubiquitin survives the shredder
What catalyzes the transfer of amino groups from AA’s to Alpha-ketoglutarate?
What is the major co-enzyme in this rxn?
Aminotransferases
Pyridoxal Phosphate
What catalyzes the transfer of an amino group of Alanine to Alpha-ketoglutarate to form pyruvate and glutamate?
Alanine > Pyruvate
A-ketoglutarate > Glutamate
Alanine Aminotransferase (ALT)
THE RULE
remember: co-enzyme is pyridoxal phosphate
Glutamate > Alpha-Ketoglutarate
Oxaloacetate > Aspartate
amino goes from glutamate to aspartate. What is this called?
What is the co-enzyme?
Aspartate aminotransferase
Pyridoxal phosphate
What are 3 important reactants and products of aminotransferase (deamination)?
Glutamate > Alpha-Ketoglutarate
Alanine > Pyruvate
Aspartate > oxaloacetate
What AA spontaneously goes through oxidative deamination?
Where does this occur?
What enzyme is involved?
What does this produce?
Why are ATP and GTP allosteric inhibitors of this rxn?
Glutamate Liver (mitochondria) Glutamate dehydrogenase free ammonia (and alpha-keto acid) NADH is produced (AMP activates)
What is the key controlling regulatory step in the urea cycle?
What allosterically regulates?
What is the product?
Where does the amino group come from?
CPSI (Carbamoyl Phosphate Synthetase I)
N-acetylglutamate (high concentrations after eating)
Carbamoyl Phosphate
ammonia comes from Glutamate
Where in the body does the urea cycle occur?
Where in the cell?
Liver (principally)
2 rxns in mitochondria (including key CPSI/Carbamoyl-P/N-acetylglutamate rxn)
rest in cytosol
Where does the Nitrogen in urea come from?
How many ATP are required to form urea?
Glutamate and Aspartate
4
What is the benign way we transport ammonia in blood?
via Alanine and Glutamine
What is a product of the urea cycle, an intermediate in the CAC, and can make aspartate in an aminotranferase (transamination) reaction with oxaloacetate?
also a link to GNG
Fumarate
What does glutaminase do?
Glutamine > Glutamate
in the liver
What happens if the glutamate dehydrogenase reaction is pushed too far toward the formation of glutamate?
It depletes Alpha-Ketoglutarate, which is essential for the CAC to run
(remember Alpha-ketoglutarate + NH3 = glutamate)
this is ammonia toxicity
What are the 2 kinds of hyperammonemia?
Acquired (liver cirrhosis)
Hereditary (mental retardation usually occurs)
What transfers the most reduced (methyl) groups?
S-adenosyl methionine
What transfers intermediately reduced carbon groups?
Tetrahydrofolate
Is CO2 considered part of the one carbon pool?
How is it transferred?
NO
Biotin (vitamin B7)
Where does folic acid come from?
What is its structure?
Vitamin B9
pteridine ring, p-aminobenzoic acid (PABA), glutamate
only bacteria make the link between pteridine and PABA
What is ingested folic acid metabolized to?
What steps and enzyme involved?
Tetrahydrofolate (THF)
dihydrofoltate reductase
two reducing steps (using two NADPH)
Folic acid rxn:
Folic acid > dihydrofolate (NADPH>NADP+) > tetrahydrofolate (NADPH>NADP+)
What drug is an analog of dihydrofolate and important in cancer treatment?
What does it inhibit?
Methotrexate
enzyme of rxn: dihydrofolate reductase
What is the most common vitamin deficiency in the world?
Folate
What 3 molecules is tetrahydrofolate a requirement to metabolize?
Purine, Thymine, and AA metabolism
Where does THF usually get its carbon from?
this is the major source of carried carbon
Serine
serine is converted to glycine in the process
What rxn forms S adenosyl methionine (SAM)?
What catalyzes?
methionine + ATP
methionine adenosyl transferase
What does SAM carry?
ONE thing.
A methyl group
How is methionine formed?
enzyme?
Homocysteine + CH3 (from THF) > Methionine
methyltransferase
What is the rate limiting step of the active methyl cycle?
MTHFR
Methylene tetrahydrofolate reductase
Summarize the activated methyl cycle:
Methionine+ATP=SAM (methionine adenyl transferase) >
SAM donates methyl group (methyl transferase) >
SAHC (- adenosine by hydrolysis) >
Homocysteine methylated by methyl-THF (methionine synthase) >
Methionine
What provides the methyl-THF for the Homocysteine>methionine step in the methyl cycle?
What’s the motherfuckin enzyme for this rxn?
Methyl THF
MTHFR
What creates the methyl trap?
What accumulates?
methyl-THF, once made, is irreversible
methyl FH4 (if not accepted by homocysteine)
What are 2 conditionally essential amino acids?
Tyrosine (from phenylalanine)
Cysteine (from homocesteine + serine)
Pyruvate + amine =
Oxaloacetate + amine =
Alanine
Aspartate
Alpha-ketoglutarate + amine =
What is the enzyme converting product to glutamine?
Glutamate
Glutamine synthetase
What is the pathway that produces cysteine and glycine?
3-Phosphoglycerate > Serine, then
- with 1C transfer to THF via MTHFR > glycine
- homocysteine > cysteine
(remember, homocysteine comes from methionine, an essential AA)
What important molecule is made from cysteine?
Glutathione
rids ROS in H2O2 pathway
How is tyrosine made? What enzyme (a deficiency in which causes PKU) is involved?
Phenylalanine >
phenylalanine hydroxylase
Tyrosine
What causes PKU?
autosomal recessive
phenylalanine hydroxylase dysfunctional
(phenylalanine cannot metabolize to tyrosine)
Where is Heme made and what is the limiting step?
Liver and bone marrow
DALA synthase is rate limiting step
What is the must-know step in Heme synthesis?
enzyme?
Where does it occur?
Succinyl CoA + Glycine > Delta aminolevulinate (DALA)
DALA synthase
mitochondria of liver and bone marrow
What is the coenzyme used for Heme synthesis and how is Heme production regulated?
pyridoxal phosphate (vitamin B6)
allosterically regulated by Heme
Myoglobin consists of a single _____, while hemoglobin consists of two ____.
polypeptide chain with 8 alpha helices
alpha/beta dimers
What is special about the Hba1c type of hemoglobin?
binds glucose non-enzymatically
How many heme groups are found on hemoglobin?
myoglobin?
4
1
Hemoglobin is most likely to accept oxygen in what conformation?
give oxygen?
Relaxed
T (taut)
What stabilizes the T-conformation of hemoglobin?
What does this do?
2,3 bisphosphoglycerate (a byproduct of glycolysis)
this makes it more likely hemoglobin will give up oxygen and deliver it to the tissues that need it the most. (airplane looking at smokestacks)
T/F
Over the physiological range of blood, myoglobin is usually saturated with oxygen.
True
What are the 3 main markers of metabolic industry that allosterically affect hemoglobin?
H+
CO2
2,3- bisphosphoglycerate
T/F
2,3 bisphosphoglycerate is released at the lungs
False
it is released before the lungs (only in smokers is the statement true)
What is the Bohr effect?
additive allosteric effects of H+ and CO2
CO2 reacts with terminal amine group on hemoglobin that forms _______ and stabilizes the T conformation.
Carbamate
carbamate is also a CO2 carrier to the lungs
What are 4 allosteric effectors of hemoglobin?
O2
H+
CO2
2,3 bisphosphoglycerate
How does carbon monoxide out compete oxygen?
Hemoglobin has a 200x affinity for CO
How does fetal hemoglobin differ from adult?
Fetal Hb has higher O2 affinity
does not bing 2,3 BPG
What accounts for about 14% of CO2 transport to the lungs?
carbamate
RBC transport - rest goes as bicarbonate
What is the major form of CO2 transport to the lungs?
What enzyme is involved?
CO2 to bicarbonate (HCO3-)
carbonic anhydrase (Zn)
What AA substitution creates sickle cell trait?
Valine for a glutamate in beta globin at 6th position
glutamate kicks out valine - this is a polar for a nonpolar AA
Does sickle cell train affect oxyhemoglobin state?
no
affects deoxy solubility
Nucleoside refers to:
Sugar and Base
What is the difference between deoxyribonucleotide and ribonucleotide?
Both are nitrogenous base (AGCTU), ribose sugar, phosphate
Deoxy is dehydroxylated at the C2 position in ribose
What is the difference between the salvage and De Novo pathways?
salvage - recycles
de novo - PPP > PRPP (activated ribose)
*de novo is expensive
What are the purines?
Pyrimidines?
A and G (two rings)
T, U, C
What does a nucleoside lack to be a nucleotide?
phosphate
Name 4 precursors for purine
he would hate to ask this question
CO2, glutamine, aspartate, glycine
need PRPP
What are the precursors for Pyrimidine?
Aspartate, glutamine, CO2
need PRPP
What is PRPP and how is it made?
phosphorylated x2 Ribose 5-P (from PPP)
PRPP synthetase
What is the first committed step in de novo purine biosynthesis?
What allosterically inhibits/activates?
PRPP > 5-Phosphorylribosyl-1-amine
via PRPP amidotransferase
allosterics:
ATP, ADP, AMPTP, GDP, GMP (different sites) inhibit (because purines)
PRPP activates
What is the function of PRPP?
Provides the activated sugar for synth
How do purine and pyrimidine synth from PRPP differ?
Purine builds onto sugar
Pyrimidine builds base and adds to sugar
What are the precursors and the committed step in de novo pyrimidine synthesis?
Aspartate, Glutamine, CO2
CPS II (carbamoyl phosphate synthetase II)
What are the 2 steps of de novo pyrimidine synth?
Carbamoyl formation
OMP
How is carbamoyl formed
remember: this is step 1 of de novo pyrimidine synth
2 ATP + bicarbonate + glutamine (for nitrogen) > Carbamoyl phosphate
via CPSII enzyme
What does carbamoyl phosphate combine with to form a pyrimidine ring?
Aspartate
What is the enzyme carbamoyl phophatase II inhibited/activated by?
inhibited - UTP
activated - ATP and PRPP
What are the parent molecules in purine and pyrimidine synth?
purine = IMP (inosine monophosphate)
pyrimidine = OMP (orotidine-5-monophosphate)
What are the rate limiting steps for purine and pyrimidine synth?
purine:
PRPP > 5-phophoribosylamine (PRPP amidotransferase)
pyrimidine:
HCO3- + NH3 (from glutamine) +2 ATP > carbamoyl phosphate (CPSII)
How are ribonucleotides converted to deoxyribonucleotides?
Step 1: RNR - ribonucleotide reductase (dehydroxylate my C2)
Step 2: thioredoxin (reduces OH)
then, NADPH regenerates thioredoxin via thioredoxin reductase
Urate causes gout. What is its soluble form?
Hypoxanthine
How does the Purine salvage pathway work?
Combine a base with PRPP
What are the two purine salvage pathways?
APRT: Adenine phosphorybosyl transferase (adenine + PRPP)
HGPRT: Hypoxanthine-Guanine phosphorybosyl transferase
Why is Pyrimidine salvage rare?
What enzyme is used if they are salvaged?
They usually degrade and excrete
Pyrimidine phosphoribosyltransferase
After pyrimidine phosphoribosyltransferase catalyzes PRPP onto a base, what is the next step?
U, T, and C are phosphorylated by their respective kinases
What is defective in Lesch Nyhan syndrome?
HGPRT enzyme (can’t recycle purines)
this causes gout from too much uric acid
5 stages of cell cycle:
G0 - resting G1 - growing (11 hrs) S - DNA replication (8 hrs) G2 - continued growth (4 hrs) M - division
What’s in a nucleosome?
8 Histone proteins
around each protein is wrapped 146 bp (1.75 turns)
T/F
In every instance, DNA replication requires an RNA primer.
True
How does the lagging strand replicate?
Still 5’ to 3’, but in Okazaki fragments
What is the function of DNA polymerase?
Catalyzes the formation of the sugar phosphate (backbone) connection in DNA
What are the 3 requirements for DNA polymerase to synthesize DNA?
- ATCG (nucleotide triphosphates present)
- Template strand
- Primer
How big is the RNA primer and why is it needed?
10-20 nucleotides long
DNA polymerase can’t initiate (but can continue)
What is the RNA primer made from?
Does it incorporate itself into DNA?
DNA polymerase alpha
No. It’s removed and replaced after the strand has fully replicated
How many RNA primers does the leading strand have?
Lagging strand?
1
multiple (each okazaki fragment)
Why might it be better to be the lagging strand in replication?
Lagging strand doesn’t get shortened (Okazaki’s repair themselves at each fragmentation)
Leading strand doesn’t replace primer with DNA. SHORTENS (only 50-80 mitotic divisions possible)
The primase/DNA polymerase use _____, but incorporate ______.
Triphosphates
Monophosphates
(attached to the pentose sugar)
What splices together Okazaki fragments?
DNA ligase
What are the terms Processivity and Fidelity mean pertaining DNA synthesis?
Processivity - enzyme ability to catalyze consecutive rxns (50k nucleotides)
Fidelity - very low error rate
How is high processivity attained in DNA synth?
Through RFC and PCNA (clamping mechanism)
What creates the replication fork? (two enzymes)
DNA helicase opens strand
Topoisomerase uncoils
Summarize leading strand synthesis:
RNA primer RFC, PCNA, DNA Polymerase bind nucleotides added, sliding down from 5' to 3' RNA primer leaves hole Telomerase (in some cases) repairs gap
Summarize lagging strand synth:
Multiple RNA primers
RFC, PCNA, DNA Polymerase
RNA primers leave and replaced by DNA polymerase
DNA ligase joins fragments
What is the only enzyme able to work with nucleoside monophosphates?
DNA ligase
What clinical technique uses palindromic sequences?
Restriction endonuclease
Blots: Southern Northern Western Eastern
DNA
RNA
Proteins
trick exam question
What are the 3 layers that ensure DNA replication accuracy?
Intrinsic fidelity (1/1000)
Proofreading (1/10 mil)
scanning enzymes/repair enzymes(1/1 billion)
What are the enzymes responsible for the immediate “proof reading” of DNA?
DNA polymerase makes error, stops
Exonuclease repairs
note: exonucleases go in both directions
What errors result in the following: single replacement stop codon protein missing all or part of exon sequence shifting sequence
missense
nonsense
RNA-splicing mutation
frameshift mutation
What is a trinucleotide expansion?
Mutation 3 bp’s repeat
What are the 5 forms of DNA damage discussed in lecture?
Mispairing (wrong bases line up)
Depurination (A/G fall out)
Deamination (creates confusion in replication)
Chemical damage
UV damage (dimers, covalent bonds, nicks)
The addition of a methyl group to guanine (via chemical damage) causes it to pair with what (instead of cytosine)?
Thymine
UV light, in particular, causes damage in the form of covalent bonding in what dimer?
TT
(thymines) 90% of UV damage
What is the enzyme used in the Direct Reversal of methylated Guanine?
O6-methyguanine methyltransferase
What is the main enzyme used in the very specific type (single base) excision repair?
Gycosylase
What 3 steps (4 enzymes) are used in the excision repair of longer strips of DNA?
Nuclease
DNA Helicase
DNA Polymerase + DNA Ligase
When can double-stranded DNA breaks be repaired and what process utilizes this?
During replication
Homologous/non-homologous recombination
A defect in the nucleotide excision repair system leads to what pathology?
Xeroderma Pigmentosum
What is the major watchman of the cell (defect of which is found in 50% of cancer patients)?
What does this watchman normally do?
p53 pathway
programs cell apoptosis
What is the class of eicosanoid derived from Omega 6 linoleic acid?
Arachidonic Acid
What is the class of eicosonoid derived from Omega 3 linolenic acid?
Eicosapentanoic acid (EPA)
What end is the modified G nucleotide attached to in RNA?
What end are the A nucleotides attached to in RNA?
5’ (G)
3’ (AAAA’s)
What is the process of exon shuffling to make RNA strands (and therefore different proteins)?
Alternative splicing
What is a promoter?
What are two gene sequence features?
A region of DNA that initiates transcription of a certain gene.
TATA box and CAAT box (sometimes GC)
What does hnRNA signify?
The cap site (PyAPy)
What does the protein start and stop signal look like on DNA/RNA?
ATG - TGA (DNA)
AUG - UGA (RNA)
Describe mRNA from 5’ to 3’ (anatomy).
5’ cap - AUG - (coding sequence) - UGA - poly A tail - 3’
What sequence denotes the cap site (where transcription begins)?
ACATTTG
The promoter region, consisting of the TATA (or CAAT) box, is responsible for binding what?
RNA polymerase
What are the two areas found within promoter sites in eukaryotes?
How big is the promoter site?
CAAT and GC are the upstream activating sequences
TATA box
100 bp long
If RNA polymerase is the helicopter, the promoter is the…
helipad
What determines the specificity of transcription of a certain gene?
the Promoter
How many types of RNA polymerase are there in eukaryotes?
3
What is the name of a family of transcription factors?
C/EBP’s (CCAAT-enhancer)
What do enhancers do?
open up chromatin, recruit other enhancers
And physically contacts promoter
What does the Leader Sequence refer to?
the 50 bp’s preceding the ATG start codon
What is the translation termination codon?
What follows it?
TAA
poly A tail
Are promoters transcribed?
No. they designate where transcription begins and are just upstream
What binds the promoter sequence before RNA polymerase can bind?
Transcription factors
What is the basal transcription apparatus?
The promoter, transcription factors, and RNA pol II acting together
What proteins interact with the basal transcription apparatus to regulate the transcription rate?
Activator proteins
What type of RNA stays in the nucleus and regulates other RNA?
snRNA
What is the mediating factor between RNA pol II and promoter sites?
Transcription factors
What direction does RNA polymerase travel?
5’ to 3’
Does RNA polymerase require a primer?
no
T/F
RNA polymerase unwinds, transcribes, and rewinds DNA
True
When does the elongation phase of transcription end?
When termination signal reached
What does the 5’ cap and the 3’ tail consist of?
methylated guanosine
200 A tail
Where is RNA pol I found?
nucleolus
Which RNA is the major one in mammals?
rRNA (85%)
What 3 sites must be in place for proper splicing and removal of introns?
5’ splice site (GUA)
Branch site (A)
3’ splice site (AG)
What is the function of snRNP’s?
bind introns (at 3 sites)
What is the structure of tRNA
coverleaf (secondary H-bond structure)
covalent ester bond to AA
anti-codon
What do all tRNA have at their 3’ terminus?
CCA - then ester bond to AA
5’ doesn’t connect to anything
How many nucleotides long is tRNA
70-80
What binds tRNA to Amino Acids?
the matchmaker
Aminoacyl tRNA synthetase
How many aminoacyl tRNA synthetases are there?
20
What are the two components of the ribosome?
What do they make when combined?
40S and 60S
80S
What ribosome binds the mRNA codon
40S
What does the tRNA capable of initiating translation carry?
methionine
What recognizes met-tRNA and binds it to the 40S ribosomal unit?
note: at least 10 of these are required to initiate translation
Eukaryotic initiation factor 2
During initiation of translation, what attaches to the methyl-guanosine cap at the 5’ end?
Cap binding proteins
eukaryotic factors 4A and 4B
What needs to occur before the 60S ribosomal subunit binds in translation?
The 40S needs to travel down to the AUG start codon
AUG is start codon. What is start anticodon?
CAU
What are the 3 binding sites in a ribosome?
A (aminoacyl)
P (peptidyl)
E (Exit)
What brings the tRNA to the A-site?
like an usher
EF-Tu
with GTP
What regenerates the spend EF-Tu GDP after ushering a tRNA?
EF-Ts
What are the 3 stop codons and what recognizes these?
UAA UAG UGA
Release factors
What do release factors bind to?
The termination codon
UAA UAG UGA
Where does the unusual base pairing occur according to the wobble hypothesis?
3rd base codon (1st base anticodon)
variability occurs in 3rd base codon
What is the function of Ferritin?
binds iron
What happens to Ferritin if iron is scarce?
IRE-BP binds IRE (Iron Response Element), blocking translation of Ferritin
