Describe three effects that account for the thermodynamic driving force for the hydrolysis of ATP (phospho

Chemistry 451

Quiz 3 (25 pts; 2 points each unless indicated)

Place each of the characteristics listed on the left under fatty acid (b ) oxidation or fatty acid biosynthesis:

Characteristic	b oxidation	Fatty Acid Biosynthesis
Occurs in mitochondrion Occurs in cytoplasm	Occurs in mitochondrion	Occurs in cytoplasm
ACP is acyl group carrier CoA is acyl group carrier	CoA is acyl group carrier	ACP is acyl group carrier
Coenzymes are NADPH Coenzymes are FADH₂and NADH	Coenzymes are FADH₂and NADH	Coenzymes are NADPH
Hydration/dehydration involves L b -hydroxyacyl group Hydration/dehydration involves D b -hydroxyacyl group	For example: L b -hydroxyacyl group	D b -hydroxyacyl group
C₂ unit product/donor is acetyl- CoA C₂ unit product/donor is malonyl-CoA	C₂ unit product/donor is acetyl- CoA	C₂ unit product/donor is malonyl-CoA

What is an "acyl group carrier"?

An acyl group carrier is a molecule like coenzyme A or carnitine or acyl carrier protein that forms a bond with the carboxyl group that can be easily hydrolyzed (typically a thioester bond). It acts to transfer the fatty acyl group between molecules. Carnitine is a carrier molecule that also acts to transport the acyl group across the inner mitochondrion membrane.

Some of us put on a few pounds this time of year because we eat more and exercise less. Acetyl CoA, the starting material for fatty acid synthesis, is generated in the mitochondrion by the oxidative decarboxylation of pyruvate (catalyzed by pyruvate dehydrogenase).

What role does the tricarboxylate transport system play in fatty acid biosynthesis?

The tricarboxylate transport system effectively transports the acetyl group on acetyl CoA (in the form of citrate) from the mitrochondrial matrix to the cytosol.

In the matrix, citrate synthase catalyzes transfer of the acetyl group on acetyl CoA to oxaloacetate to form citrate. A transporter protein then transports citrate across the inner mitochondrial membrane. In the cytosol, citrate lyase catalyzes transfer of the acetyl group from citrate to CoA forming acetyl CoA and oxaloacetate. See page 583

If you blocked the tricarboxylate transport system what effect do you think it would have on fatty acid biosynthesis?

Fatty acid synthesis should slow down because acetyl CoA would not be transported from the mito to the cytosol where fatty acid biosynthesis occurs.

The opposing pathways of fatty acid degradation and synthesis are hormonally regulated. Glucagon (released when blood glucose is low; i.e.,when you are hungry) and epinephrine (the hormone that mediates the fight/flight response) activate hormone sensitive lipase and inactivate acetyl-CoA carboxylase.

a. Name or draw the general structure of the fuel that is hydrolyzed by hormone sensitive lipase AND two products formed in the reaction catalyzed by hormone sensitive lipase (or pancreatic lipase)?

Triglyceride ® glycerol + 3 fatty acids.

(Technically triglyceride ® 2 fatty acids + 2-acyl glycerol; pg. 564, but you get the idea)

b. Name or draw the structure of the product necessary for fatty acid synthesis that is formed in the reaction catalyzed by acetyl-CoA carboxylase?

Malonyl CoA

Given the role of hormones in regulation of fatty acid metabolism which of the following mechanisms would you expect to play the greatest role in regulation of enzymes involved in fatty acid metabolism?

a. Covalent modification	b. Substrate Cycling	c. Allosteric Effects	d. Genetic Control (enzyme induction)
e.g. phosphorylation/dephosphorylation

Evidence suggests that malonyl CoA inhibits appetite. Assuming you like to invest in the stock market (and the Supreme Court acts rationally), would you buy stock in a company that planned to develop fatty acid synthase inhibitors as diet pills? Why or why not? (I'm not looking for anything specific here - just give an opinion)

I'd buy… But, the Supreme Court blew it…

Nitrogen is either moved around between amino acids or eliminated as urea. Identify the following examples as transamination or oxidative deamination.

Reaction	Transamination OR Oxidative Deamination
Aspartate ® Oxaloacetate	Transamination
Glutamate + NAD⁺ ® a -ketoglutarate + NADH + NH₃	Oxidative Deamination
Alanine ® Pyruvate	Transamination
Glutamate ® a -ketoglutarate	Transamination

Which process, transamination or oxidative deamination results in net deamination (i.e., release of free ammonia)? (1 pt)

oxidative deamination

Draw the general structure of an a -amino acid and corresponding a -keto acid.

See structures on pg. 616

Briefly, discuss or diagram what you like best about the urea cycle.

My favorite aspect of the urea cycle is that CPS-I catalyzes the same reaction as seen in the first step in pyrimidine biosynthesis (catalyzed by CPS-II).

Answer ONE of the following (a or b):

Explain why a low glucose affinity (high Km of ~5mM) for glucokinase allows glucokinase activity to increase rapidly with blood [glucose]. Why can hexokinase (with a Km<0.1mM) not respond rapidly to a change in blood [glucose] from 5 to 6 mM?

Low affinity for glucose means that even at high glucose concentrations (5 mM) glucokinase will not be saturated. This means rate of phosphorylation will increase with increasing [glucose]. In contrast, at 5 mM [glucose] hexokinase is saturated so rate of glucose phosphorylation will be constant and will not increase with increasing [glucose]. Importantly, once glucose is phosphorylated, it is trapped inside the liver cell. In this way, glucokinase acts to buffer plasma concentrations of [glucose].

Insulin promotes translocation of GLUT 4 to the plasma membrane. Why does increasing the number of GLUT 4 transporters increases the Vmax of glucose transport into muscle and adipose tissue where GLUT 4 transporters are expressed?

Rate of transport is directly proportional to the number of transporters.

Describe a metabolic defect associated with impaired purine metabolism or elimination of a purine metabolite.

Lesch-Nyhan syndrome (pg 700) results from a severe HGPRT deficiency. (HGPRT catalyzes the transfer of ribose-5-phosphate from PRPP to hypoxanthine and guanine to form IMP and GMP). Without HGPRT, PRPP builds up and because of feed forward activation of reaction #2 in purine synthesis, accelerates synthesis of purines. Increased levels of purines leads to an increase in the purine degradation product (uric acid). Lesch-Nyhan syndrome includes neurological abnormalities such as spasticity, mental retardation, and highly aggressive and distructive behavior, including self mutilation. The mechanism responsible for the neurological symptoms is not understood.

Orotic aciduria (pg 704) results from a deficiency in the enzyme that catalyze reactions 5 and 6 in de novo synthesis of pyrimidines. An excess of orotate is present in the urine. Symptoms include suppressed growth and severe anemia.

Severe combined immunodeficiency disease (SCID) (pg 715) results from deficits in the gene coding for adenosine deaminase. Adenosine deaminase removes the N in adenosine and deoxyadenosine to form inosine and deoxyinosine. Deficient adenosine deaminase causes an accumulation of deoxyadenosine and subsequently elevated dATP. High [dATP] inhibits ribonucleotide reductase (the enzyme that removes the OH on C₂’ to form deoxynucleotides from nucleotides). Inhibition of ribonucleotide reductase prevents the synthesis of other dNTPs, choking off DNA synthesis and thus cell proliferation. Lymphocytes are especially susceptible because lymphocytes produce a lot of deoxyadenosine which then builds up in the absence of adenosine deaminase. SCID is a fatal genetic disorder that has led to failed as well as successful attempts at gene therapy. (See Anderson W.F., 2000, The Best of Times, the Worst of Times, Science, 288(5466), pg 627.)

Gout (pg 717) is caused by an excess of uric acid, the final product of purine metabolism. The most common cause of gout is impaired uric acid excretion. Symptoms include inflammation of the joints and kidney stones.

Annotate ONE of the following three figures to describe the signal transduction cascade beginning with ligand binding and ending with cellular response.

Fig. 21-12; pg. 674-677 (See Guided Exploration 19: Mechanisms of hormone signaling involving the Adenylate Cyclase system)

Ligand binds to stimulatory receptor (Rs).
Binding of ligand causes GDP to dissociate and GTP to bind to stimulatory alpha subunit (s-alpha) of heterotrimeric G protein
Once GTP binds, s-alpha subunit dissociates from gamma-beta subunit.
s-alpha subunit diffuses laterally in membrane until it bumps into adenylate cyclase.
s-alpha activates adenylate cyclase which then catalyzes formation of cAMP from ATP.
cAMP then binds to and activates cAMP dependent protein kinase (R2C2). The activated kinase (2C) then phosphorylates a protein which leads to a cellular response.
cAMP is hydrolyzed (and inactivated) by phosphodiesterase.
s-alpha continues to activate adenylate cyclase until GTP is hydrolyzed to GDP.
Continued occupancy of the receptor by its ligand, results in repeated cycles of G-protein activation and hence a prolonged cellular response.
Ligand binding to an inhibitory receptor (Ri) produces a similar cascade, however, the inhibitory alpha subunit inhibits adenylate cyclase until GTP is hydrolyzed to GDP which turns off the inhibitory alpha subunit.

Fig. 21-16; pg. 677-682 (See Guided Exploration 20: Mechanisms of Hormone Signaling Involving the Receptor Tyrosine Kinase System AND Interactive Exercises for Fig. 21-13).

Ligand (protein growth factor) binds to extracellular portion of dimeric receptor tyrosine kinase (RTK) which causes intracellular portion of dimers to associate (see Fig. 21-13)
Joining of the intracellular parts of the receptor (dimerization of the intracellular domains) activates the receptor (tyrosine kinase).
The activated tyrosine kinase catalyzes phosphorylation of itself so that tyrosine residues (Y) are phosphorylated.
SH2 is a common domain on many cytoplasmic proteins that bind to phosphorylated receptors. When SH2 binds to the phosphorylated receptor another part of the protein (the SH3 domain) binds to Sos (another protein) forming a complex that bridges the activated receptor tyrosine kinase with a GTP binding protein called Ras.
Ras is activated by GTP binding (GDP dissociates)
Activated Ras binds to and activates Raf, which phosphorylates MEK, which phosphorylates MAPK
MAPK moves into the nucleus and phophorylates a bunch of transcription factors like Myc, Fos and Jun which turn on gene expression…

(Why do they call it a kinase cascade?)

Fig. 21-19; pg. 683-685: The Phosphoinoside Pathway

Note: Phosphatidylinositol-4-5-bisphosphate (PIP2) is a phospholipid that is found in the plasma membrane (inner leaflet). Phospholipase C catalyzed hydrolysis of PIP₂ produces diacylglycerol (DG) and Inositol-1,4,5-trisphosphosphate (IP₃). DG remains in the membrane. IP₃ is a water soluble sugar that diffuses into the cytoplasm

Ligand binds to extracellular domain of receptor
Activation (change in conformation) of receptor causes GTP to bind and GDP to dissociate.
Once GTP binds, q-alpha subunit dissociates from gamma-beta subunits.
q-alpha subunit diffuses laterally in membrane until it bumps into phospholipase C (PLC).
q-alpha activates PLC which catalyzes hydrolysis of PIP₂ to form DG and IP₃.
IP₃then binds to and activates an IP₃ gated Ca²⁺ channel in the endoplasmic reticulum membrane.
Ca²⁺ binds to calmodulin and activates calmodulin-dependent protein kinase.
The activated protein kinase phosphorylates proteins that lead to a cellular response.
DG stays in the membrane where, with the help of Ca²⁺ , activates Calcium dependent protein kinase (PKC). This kinase phosphorylates proteins that lead to a cellular response.
q-alpha continues to activate PLC until GTP is hydrolyzed to GDP.
Continued occupancy of the receptor by its ligand, results in repeated cycles of G-protein activation and hence a prolonged cellular response.