High Yield Topics: Biochemistry—Metabolism

Glycogen sythesis: regulation and inherited defects.

· Glycogen is the storage form of glucose, mainly found in liver and muscles.

· The synthesis and breakdown of glycogen are occurring constantly by the body.

· The breakdown of gylcogen to glucose-1-P and glycogen chain (with one less glucose) is catalyzed by Phosphroylase in the liver. Phosphorylase breaks the a(1®4) bond between the glucose units in gylcogen.

· For the Synthesis of glycogen, UDP-glucose is needed. The pathway is:

Glucose ® Glucose-6-P --(via phosphoglucomutase)® glucose-1-P + UTP | (pyro- ¯phosphorylase)

Glycogen chain (n+1) + UDP ¬(Glycogen Synthase)-- UDP glucose + PPi

· Glycogen Synthase is the enzyme that attaches glucose to the tyrosine group of the glycogen chain fragment. If no chain is available, glucose is attached to glycogenin (a protein that can accept the glucose and start a new glycogen chain). Synthase only makes the a(1®4) bond between the glucose units, so there is a branching enzyme that makes the a(1®6) bond to make branching glycogen chains.

· Regulation of Glycogen Synthesis:

o Phosphorylation regulates the activity of Glycogen Synthase (GS): when GS is phosphorylated, it is Inactive. When it is dephosphorylated (by a phosphatase enzyme), it is Active. It can also be allosterically activated by Glucose-6-P, in a feed-forward mechanism. (with the G-6-P activation, the GS is still phosphorylated, yet it is active!).

o Insulin activates GS by producing phosphatase activity. This makes sense since Insulin promotes glycogen synthesis.

o Inhibitors of GS include: Phosphorylase (in liver)—this is activated by Glucagon and Epinephrine (via cAMP and PKA transduction). In muscle, cAMP and Calcium inhibit GS by turning on phosphorylase. It makes sense that Glucagon and Epinephrine will inhibit GS because they want to decrease glycogen synthesis and promote gluconeogenesis (release of glucose).

o Phosphatase (induced by insulin) can actually inhibit phosphorylase activity to increase glycogen synthesis.

· Glycogen Storage Diseases (12 total) are inherited defects in glycogen metabolism and cause abnormal accumulation of glycogen within the cell. The important ones are listed in First Aid pg. 146 (2000 Edition) and pg.150 (’01 ed)

Oxygen consumption, carbon dioxide production, and ATP production for fats, protein, and carbohydrates.

· Energy Yield from Fatty Acid oxidation: 131 ATP from 1 molecule of palmitoyl coA. 7 NADH and 7 FADH2 were produced along with 8 acetyl coA. For more details, see Lippencott’s Biochemistry, pg183. Oxygen consumption or Carbon dioxide production could not be found!!

· I also could not find ATP production for protein or carbos (I’m sorry!)

· For an added bonus, I’ll include a Fatty acid enzyme deficiency: Medium-chain fatty acyl CoA dehydrogenase deficiency. This is found in 1/10,000 births (more common than PKU!!). It causes a decrease in FA oxidation and severe hypoglycemia. It is responsible for up to 10% of SIDS (sudden infant death syndrome)

Amino Acid degradation: Urea Cycle and TCA cycle.

· Urea is the main nitrogen-containing component of the urine. It is made in the liver, then transported via blood to the kidneys for excretion. Urea formation occurs both in mitochondria and in cytosol.

· The urea cycle (First Aid, pg 159 for 2000 ed, pg 164 for 2001). Key points is that Carbamoyl-P Synthetase I converts CO₂ and NH₄⁺ into Carbamoyl-P, and the enzyme needs N-acetylglutamate to work. Carbamoyl-P then joins with ornithine to form citrulline (and on down the pathway).

· Aspartate provides the 2^nd nitrogen of the urea molecule during the cycle. But, Glutamate forms both ammonia and Aspartate which both enter the cycle. So glutamate is essentially contributing both nitrogens in the urea cycle!

· TCA cycle: many amino acids are degraded by conversion into other end products. Amino acids can be classified as: (from Lippincott’s)

· Ketogenic aa: Amino acids whose catabolism yields acetoacetate (ketone body) or it’s precursor (acetyl coA and acetoacetyl coA). Leucine and lysine are the only exclusively ketogenic aa found in proteins.

· Glucogenic aa: Amino acids whose catabolism yields pyruvate or a TCA intermediate both which are substrates for gluconeogenesis. Look at Lippincott’s for a list of aa and where they can enter the TCA cycle!

Effect of enzyme phosphorylation on metabolic pathways.

· Phosphorylateion is an important way for enzyme regulation of metabolic pathways. For example know the action of Glucagon and Insulin and what they do to regulate enzymes.

· Glucagon will activate Protein Kinase A (PKA) via cAMP. PKA will phosphorylate enzymes which will lead to gluconeogenesis and decrease gylcolysis.

· Insulin will activate phosphatase, which de-phosphorylates many enzymes to promote glycolysis. An example is Fructose 2,6-Bis P, which stimulates Phosphofructokinase (PFK) to make fructose 1,6-Bis P which drives the glycolysis pathway.

· Other examples include phosphorylation of Glycogen synthase for glycogen synthesis.

Rate-limiting enzymes of different metabolic pathways:

· Glycolysis and Gluconeogenesis: irreversible enzymes found in first aid. (pg 154 for 2000 ed)

· Other Regulated/rate-limiting enzymes :

Pathway	Enzyme
FA synthesis	ACC = acetyl coA carboxylase
Cholesterol synthesis	HMG-CoA Reductase (statins block this)
Urea Cycle	Carbamoyl-P Synthetase (makes carbamoyl-P from CO₂ and NH₄⁺
Pentose Pathway	Glucose-6-P dehydrogenase
TCA cycle	Regulated steps: Pyruvate dehydrogenase Citrate synthase Isocitrate dehydrogenase a-ketoglutarate dehydrogenase

6. Sites of different metabolic pathways (What organ? Where in cell?)

(info from pp. 156& 158 First Aid; other details from Lippincott’s Biochem)

Pathway	Metabolism Site	Organ (s)
Beta-oxidation	Mitochondria	Liver, muscle, heart
Acetyl CoA production	Mitochondria	Liver, muscle, heart
TCA cycle	Mitochondria	Liver, muscle, heart
Glycolysis	Cytoplasm	All tissues (glucokinase used in liver; hexokinase all other tissues)
Fatty acid synthesis	Cytoplasm	Most in liver & lactating mammary glands (also adipose tissue & kidney)
HMP shunt	Cytoplasm	Most impt. in liver, mammary glands, & adrenal cortex (things that make fat & steroids that require NADPH)
Protein synthesis (RER)	Cytoplasm	Liver & all other tissues
Steroid synthesis (SER)	Cytoplasm	Adrenal cortex, ovaries, testes
Gluconeogenesis	Both (pyruvateàoxaloacetate in the mito; rest in the cytoplasm)	Liver (90%) Kidneys (10%)
Urea cycle	Both (1^st 2 rxn.’s in mito; rest in cytoplasm)	Liver
Heme synthesis	Both (1^st rxn. & last 3 rxn.’s in mito; rest in cytoplasm)	Liver & bone marrow
Ketogenesis	??	Liver

7. Fed vs. fasting state: energy forms used, direction of pathways

(see p. 161 in First Aid; also a nice discussion in Chapters 25 & 26 in Lippincott’s Biochem)

Fed

Fasting

Liver

-inc. glucose-6-phosphate

-inc. glycogen synthesis

-inc. HMP shunt

-inc. glycolysis

-decr. gluconeogenesis

-inc. fatty acid synthesis

-inc. triacylglyceride synthesis

-inc. amino acid degradation

-inc. protein synthesis

-inc. glycogen breakdown

-inc. gluconeogenesis

-inc. fatty acid oxidation

-inc. ketone bodies

Adipose tissue

-inc. glucose transport

-inc. glycolysis

-incr. HMP shunt

-inc. fatty acid synthesis

-inc. triacylglyceride synthesis

-decr. triacylglyceride degradation

-inc. triacylglyceride degradation

-inc. fatty acid release

-decr. fatty acid uptake

Skeletal muscle

-inc. glucose transport

-inc. glycogen synthesis

-glucose used for energy, not fat

-inc. protein synthesis

-inc. branched chain amino acid uptake

-glucose uptake decreased

-oxidizes fatty acids for fuel

-inc. protein breakdown to make glucose precursors

Brain

-glucose used exclusively for fuel

-fat is not important

-glucose used during 1^st few days

-after 2-3 wks., ketone bodies can be used

8. Tyr kinases and their effects on metabolic pathways

(info from http://www.ncbi.nlm.nih.gov/books/mboc/mboc.cgi?code=150303480625145)

Some receptors are linked to intracellular tyrosine kinase domains. When a ligand binds to these receptors, the receptors transfer a phosphate group from ATP to certain tyrosine side chains both on the receptor and other specific cellular proteins. Receptors that are linked to tyrosine kinase domains include epidermal growth factor receptor (EGFR), platelet-derived growth factor (PDGF), fibroblast growth factors (FGFs), hepatocyte growth factor (HGF), insulin, insulin-like growth factor-1 (IGF-1), nerve growth factor (NGF), vascular endothelial growth factor (VEGF), and macrophage colony stimulating factor (M-CSF).

When the ligand(s) bind, these receptors dimerize and cross-phosphorylate one another. Sometimes the ligand binding actually brings 2 monomers together (PDGF). In other cases, ligand binding facilitates a conformational change that leads to dimerization of receptors (EDF). The insulin receptors are actually tetramers to start with and ligand binding is thought to induce an allosteric interaction of the two receptor halves rather than receptor dimerization. The exact proteins that are phosphorylated and the downstream events that occur depend on the receptor type.

9. Anti-insulin hormones (glucagon, GH, cortisol)

The explanation about these hormones could go on & on. For a nice discussion of glucagon, see pp. 275-276 in Lippincott’s Biochem. For info on GH & cortisol, see pp. 277-278 in Lippincott’s Biochem.

10. Synthesis and Metabolism of Neurotransmitters

I included only what I could find. I included the cofactors used in each reaction if I could find them. If a cofactor is not listed, it doesn’t mean that it is not involved in that reaction. I bolded the things that are probably important. Everything else is just in case you want to know more in depth. If there are any questions contact me (Bryan Holcomb) at holcombb@umich.edu.

I. Tyrosine derived neurotransmitters

A. Dopamine

1. Synthesis: Tyr --------(a)----------à L-Dopa-----------(b)---------àDopamine

a. enz. = Tyrosine Hydroxylase, cofactors = tetrahydrobiopterin and O2 gives off dihydrobiopterin and H2O.

b. enz = DOPA hydroxylase (aromatic amino acid decarboxylase), cofactors = pyridoxyl phosphate (PLP or vit. B6), gives of CO2.

2. Metabolism:

a. Major mechanism = Reuptake

b. Breakdown: dopamine ---(i)--àDHPA ----(ii)-àDOPAC----(iii)-àHVA or dopamine---(iii)à 3-methoxytyramine---(i)--àMHPA---(ii)à HVA

i. enz. = monoamine oxidase (MAO), cofactor = FAD

ii. enz. = aldehyde dehydrogenase

iii. enz. = catecholamine-O-methyltransferase (COMT), cofactor = Mg²⁺, S-adenosylmethionine (SAM)

B. Norepinephrine

1. Synthesis: Dopamine ----(a)---ànorepinephrine

a. enz. = dopamine-b-hydroxylase, cofactor = ascorbate (vit. C) and O2 give off H2O and dehydroascorbate.

2. Metabolism:

a. Reuptake is most important

b. Breakdown: norepinephrine ---(i)ànormetanephrine---(ii)àMOPGAL---(iii)àVMA(5-hydroxymandelic acid) or MOPGAL---(iv)àMOPEG.

c. norepinephrine ---(ii)à DOPGAL---(iv)àDOPEG---(i)àMOPEG

d. norepinephrine ---(ii)àDOPGAL---(iii)àDOMA---(i)àVMA

i = COMT

ii = MAO

iii = aldehyde dehydrogenase

iv = aldehyde reductase

d. Remember reuptake then COMT and MAO.

C. Epinephrine

1. synthesis: norepinephrine ---(a)à epinephrine

a. enz. = phenylethanolamine N-methyltransferase, cofactors = SAM to S-adenosylhomocysteine.

2. Metabolism:

a. epinephrine---(i)àmetanephrine---(ii)àMOPGAL (see norepinephrine for completion from this product.

b. epinephrine---(ii)àDOPGAL (see norepinephrine for completion of this pathway

i. COMT

ii. MAO

c. Basically epinephrine breaks down to the same products as norepinephrine using MAO and COMT.

Glutamate derived neurotransmitters

A. Glutamate

1. Synthesis: in axon mitochondria from the glutamine supplied by glial cells. Glutamine---(a)-àglutamate.

a. enz. = glutaminase, byproduct = ammonium

2. Metabolism:

a. axon uptake and storage

b. glial uptake and conversion to glutamine. Glutamate ---(i)àglutamine.

i. enz. = glutamine synthetase, cofactors = ATP and ammonium gives off ADP and Pi.

B. GABA

1. Synthesis: glutamate ---(a)àGABA

a. enz. = glutamate decarboxylase, cofactors = PLP, byproduct = CO2.

2. Metabolism:

a. Most important is reuptake.

b. Breakdown = GABA---(i)àsuccinic semialdehyde---(ii)àsuccinate

i. enz. GABA transaminase, cofactors = PLP and glutamate which is converted to a-ketoglutarate.

ii. Enz. = succinic semialdehyde dehydrogenase, cofactor = some oxidizing agent, probably NAD+, but I couldn’t find it.

Tryptophan (serotonin)

A. Serotonin synthesis: tryptophan---(1)à 5-hydroxytryptophan--(2)àserotonin

1. enz. = tryptophan hydroxylase, cofactors = tetrahydrobiopterin and O2 to dihydrobiopterin and H2O.

2. enz = aromatic amino acid decarboxylase, cofactor = PLP, byproduct = CO2

B. Serotonin metabolism:

1. Reuptake is important

2. Breakdown

a. Serotonin ---(i.)à5-hydroxyindoleacetic acid (5-HIAA)

b. In pineal gland: serotonin---(ii)àN-acetyl serotonin---(iii)àmelatonin

i. enz. = MAO and aldehyde dehydrogenase

ii. enz. = serotonin N-acetyltransferase

iii. enz. = 5-hydroxyindole-O-methyltransferase

Histidine (Histamine)

A. Synthesis: Histidine---(1)àhistamine

1. enz. = histidine decarboxylase, cofactor = PLP, byproduct = CO2.

B. Metabolism:

1. histamine---(a)àN-methylhistamine---(b)àN-methylimidazole acetic acid

2. histamine---(c)àimidazole acetic acid---(d)àriboside

a. enz = histamine-N-methyltransferase

b. enz = MAO

c. enz = diamine oxidase

d. several steps

Glycine and aspartate

A. Synthesis: Probably the same way that they are synthesized normally, but I could not find the exact method used in the nervous system

B. Metabolism: I could not find the method used in the nervous system.

Acetylcholine (Ach)

A. Synthesis: Acetyl-CoA + choline ---(1)àAch

1. enz. = choline acetyltransferase, byproduct = coenzyme A

B. Deactivated in the synaptic cleft by acetylcholinesterase to acetic acid and choline ( choline is reabsorbed).

Neuropeptides

A. Synthesis: all except for a few of the small ones are synthesized in the cell body and transported down the axon. (opiates are examples)

B. Deactivation: location specific enzymes.

11. Purine/pyrimidine degradation

Purines get degraded to uric acid. (see p. 349, 350 in Lippincott’s Biochem for genetic diseases associated with purine degradation)

Pyrimidines get degraded to beta-alanine (an acetyl-CoA precursor) & beta-aminoisobutyrate (a succinyl-CoA precursor).

12. Carnitine shuttle: function, inherited defects

(from p. 182 in Lippincott’s Biochem)

We need the carnitine shuttle to transport fatty acids into the mitochondria from the cytoplasm before beta-oxidation can occur. (The CoA that is attached to the fatty acid makes it so it can’t cross the mitochondrial membrane.) The transferring of acyl groups in this shuttle requires the enzymes carnitine acyltransferase I & II. If you don’t have these enzymes in your skeletal muscle, you can’t use long-chain fatty acids as metabolic fuel. You get myoglobinemia & weakness following exercise.

13. Cellular/organ effects of insulin secretion

(info below is from p. 165 in First Aid; see p. 273 in Lippincott’s Biochem for more info about the metabolic effects of insulin)

àinsulin has no effect on glucose uptake by the brain, RBC’s, or hepatocytes

àinsulin is needed for adipose & skeletal muscle to take up glucose

àinsulin inhibits glucagon release by alpha cells of the pancreas