USMLE Step 1 Review/Biochemistry

Genetics

DNA: Z vs. B form: which is inactive ZZZZ is sleeping (inactive). B form is therefore active DNA.

Cell cycle stages "Go Sally Go! Make Children!": G1 phase (Growth phase 1) S phase (DNA Synthesis) G2 phase (Growth phase 2) M phase (Mitosis) C phase (Cytokinesis)

Exon vs. intron function Exons Expressed. InTrons In Trash.

Codons: nonsense mutation "Stop talking nonsense!": Nonsense mutation causes premature stop.

Nucleotides

Nucleotides: class having the single ring• "Pyrimadines are CUT from purines" • Pyrimidines are: Cytosine Uracil Thiamine They are cut from purines so the pyrimadines must be smaller (one ring).

Nucleotides: purines vs. pyrimidines "Guardian Angels are Pure, with two Wings": G and A are Purines, with two Rings.

Nucleotides: purines "AGUA PURa": Adenine and GUAnine are PURines. • "Agua pura" is spanish for "pure water". or "PURe As Gold" Purines - Adenine - Guanine

Nucleotides: which are purines "Pure Silver": • Chemical formula of Pure silver is Ag. Therefore, Purines are Adenine and Guanine.

Pyrimidines nucleotides "CUT the PY" (cut the pie): Cytosine Uracil Thiamine are the PYrimidines

Nucleotides: double vs. triple bonded basepairs "TU bonds" (two bonds): T-A and U-A have Two bonds. G-C therefore has the three bonds.

Genetic disorders

Hurler syndrome features HURLER'S: Hepatosplenomegaly Ugly facies Recessive (AR inheritance) L-iduronidase deficiency (alpha) Eyes clouded Retarded Short/ Stubby fingers

Down syndrome features: complete "My CHILD HAS PROBLEM!": Congenital heart disease/ Cataracts Hypotonia/ Hypothyroidism Incure 5th finger/ Increased gap between 1st and 2nd toe Leukemia risk x2/ Lung problem Duodenal atresia/ Delayed development Hirshsprung's disease/ Hearing loss Alzheimer's disease/ Alantoaxial instability Squint/ Short neck Protruding tongue/ Palm crease Round face/ Rolling eye (nystagmus) Occiput flat/ Oblique eye fissure Brushfield spot/ Brachycephaly Low nasal bridge/ Language problem Epicanthic fold/ Ear folded Mental retardation/ Myoclonus

Down Syndrome- Chromosome 21/ "D"own- "D"rink at 21

Down syndrome pathology DOWN: Decreased alpha-fetoprotein and unconjugated estriol (maternal) One extra chromosome twenty-one Women of advanced age Nondisjunction during maternal meiosis

Patau's- Chromosome 13/ "P"atau's-"P"uberty at 13

Edward's-Chromosome 18/ "E"dward's-"E"lection voter at 18

DiGeorge/ Velocardiofacial syndrome: features CATCH 22: Cardiac abnormalities Abnormal facies Thymic aplasia Cleft palate Hypocalcemia 22q11 deletion

Marfan syndrome features MARFAN: Mitral valve prolapse Aortic Aneurysm Retinal detachment Fibrillin Arachnodactyly Negative Nitroprusside test (differentiates from homocystinuria)

Cri-du-chat syndrome: chromosomal deletion causing it is 5p(-) What's another name for a cat that's five letters long and starts with a P? (Answer: pussy). Why is the cat crying? Missing its P.

APKD: genetics ADult Polycystic Kidney Disease is Autosomal Dominant • Also, "Polycystic kidney" has 16 letters and is due to a defect on chromosome 16.

Hardy-Weinberg equilibrium: causes for deviations from it "Maggie May Does Not Smoke": Mutations Migration Drift Non-random mating Selection

Achrondroplasia dwarfism: inheritance pattern Achondroplasia Dwarfism is Autosomal Dominant.

Tumor suppressor vs. proto-onogene mutations: recessive vs. dominant "Recess Suppress": Tumor suppressor mutations are recessive. Proto-oncogenes are opposite (dominant).

• p53
• Retinoblastoma gene
• Breast-cancer gene
• MSH2 gene
• MLH1 gene, chromosome 3
• XP gene

Pedigree symbols: gender and affected Gender: The cIRcle is a gIRl [so boys are squares]. Affected: Black plague was a disease, so black-filled symbol means an affected/diseased person [so non-filled-in is unaffected].

Tay Sach's features SACHS: Spot in macula Ashkenazic Jews CNS degeneration Hex A deficiency Storage disease

• Extra details with TAY:
  • Testing recommended
  • Autosomal recessive/ Amaurosis
  • Young death (<4 yrs)

Chromosome 15 diseases Chromosome 15 has its own MAP: Marfan syndrome Angelman syndrome Prader-Willi syndrome

Bartter syndrome: inheritance BARtter syndrome is autosomal recessive (AR).

Imprinting diseases: Prader-Willi and Angelman "Pray to an Angel": Prader-Willi and Angelman are the 2 classic imprinting diseases. • Which disease results, depends on whether 15q deletion is maternal or paternal. Keep them straight by: Paternal is Prader-Willi. • See diagram for cardinal symptom of each disease.

Li-Fraumeni syndrome: defective p53 gene

Xeroderma pigmentosum: defective excision endonuclease

Hereditary nonpolyposis colorectal cancer: associated to mutations in MSH2 and MLH1 genes.

Mismatch repair cancer syndrome (Turcot syndrome)

Waardenburg syndrome

Useful websites

• MedMCQs: FREE Medical MCQs

Biochemistry

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Essential Amino Acids

• PriVaTe TIM HALL: Phe, Val, Thr, Trp, Ile, Met, His, Arg, Leu, Lys

=Vitamin B complex Mnemonic

Try (Thiamine:B1) Running (riboflavin : B2) not (Niacin: B3) pants (pantothenic acid:B5) partrying (pyridoxine: B6) because (Biotin :B7) fall (folic acid: B9) comes( cayanobolamine: B12).

Urea Cycle

• Ordinarily, Careless Crappers Are Also Frivolous About Urination: Ornithine, Carbamoyl, Citrulline, Aspartate, Arginosuccinate, Fumarate, Arginine, Urea.

Cell division

• "People Meet And Talk" or "PMAT": Prophase, Metaphase, Anaphase, Telophase.

In the Fasted State

• Phosphorylate Phosphorylation cascade active when blood glucose low.
• DNA expression into mature mRNA
• Exons expressed, Introns in the trash.
• Pyrimidines are CUT from purines: Pyrimidines are Cytosine, Uracil, Thiamine and are 'cut' from the 2 ringed purines, making them one ringed.

More Biochemistry

Porphyrias: acute intermittent porphyria symptoms 5 P's: Pain in abdomen Polyneuropathy Psychologial abnormalities Pink urine Precipitated by drugs (eg barbiturates, oral contraceptives, sulpha drugs)

BUN:creatinine elevation: causes ABCD: Azotremia (pre-renal) Bleeding (GI) Catabolic status Diet (high protein parenteral nutrition)

G6PD: oxidant drugs inducing hemolytic anemia AAA: Antibiotic (eg: sulfamethoxazole) Antimalarial (eg: primaquine) Antipyretics (eg: acetanilid, but not aspirin or acetaminophen)

Vitamin B3 (niacin, nicotinic acid) deficiency: pellagra The 3 D's of pellagra: Dermatitis Diarrhea Dementia • Note vitamin B3 is the 3 D's.

Tangier's disease: hallmark "Tangierene tonsils": Hallmark is large orange tonsils. • Important clinical note: there is an increased risk of atherosclerosis, just like not eating good stuff like tangerines.

Na+/K+ pump: 'K+' and 'in' each consist of 2 symbols, so 2 K+ are pumped in; 'Na+' and 'out' each consist of 3 symbols, so 3 Na+ are pumped out

Glycogen storage: Anderson's (IV) vs. Cori's (III) enzyme defect ABCD: Anderson's=Branching enzyme. Cori's=Debranching enzyme. • Otherwise, can't really distinguish clinically.

Nitric oxide: amino acid precursor When the dentist works on your teeth, you say, "AAArg! (Arginine)" before he administers Nitric Oxide (NO) to take the pain away. • Other players necessary for NO synthesis: NO synthase, Ca++, NADPH.

Pompe's disease: type "Police = Po + lys": Pompe's disease is a lysosomal storage disease (alpha 1,4 glucosidase). The POLICE (POmpe + LYSosomal) Arrest 1 of the 4 Girls (Alpha 1,4 Glucosidase)

Metabolic acidosis (normal anion-gap): causes • With hyperkalemia: RAISE K+: RTA type 4 Aldosterone or mineralocorticord deficiency Iatrogenic: NH4Cl, HCl "Stenosis": obstructive uropathy Early uremia • With hypokalemia: ReDUCE K+: Renal TA type 1 and 2 Diarrhoea Urine diversion into gut Carbonate anhydrase inhibitor Ex-hyperventilation

Galactosaemia: enzyme deficiency GALIPUT: Galactose 1 Phosphate Uridyl Transferase. • There is an assay called the Galiput test for this.

Folate deficiency: causes A FOLIC DROP: Alcoholism Folic acid antagonists Oral contraceptives Low dietary intake Infection with Giardia Celiac sprue Dilatin Relative folate deficiency Old Pregnant

Mitochondrial DNA (mt DNA) properties "mt DNA". mt stands for: Maternal Transfer Mutates Tremendously (high mutation rate)

Vitamins: which are fat soluble "The FAT cat is in the ADEK (attic)": Fat soluble vitamins are A,D,E,K.

Enzyme kinetics: competitive vs. non-competitive inhibition With Kompetitive inhibition: km increases; no change in Vmax. With Non-kompetitive inhibition: No change in km; Vmax decreases.

Glycogen storage: names of types I through VI "Viagra Pills Cause A Major Hardon": Von Gierke's Pompe's Cori's Anderson's McArdle's Her's

Sickle cell disease pathophysiology SICKle cell disease is due to a Substitution of the SICKsth amino acid of the B chain.

Glucagon function "Mr. Gluca has Gone to the cAMP to bring out some Glucose": • Glucagon elevates glucose by cAMP mechanism.

Van den Bergh reaction (Jaundice test) "Indirect reacting bilirubin = Unconjugated bilirubin": Both start with vowels, so they go together: Indirect & Unconjugated.

Hemoglobin binding curve: causes of shift to right "CADET, face right!": C = Increase in carbon dioxide, A = Acidosis (low ph), D = Increase in 2,3 DPG aka 2,3 BPG, E = Exercise, T = increase in temperature

G proteins for respective receptors "QISS & QIQ" (Kiss and Kick): • G-proteins and their respective receptors (alphabetical order): Q: alpha 1 I: alpha 2 S: beta 1 S: beta 2 & Q: M1 I: M2 Q: M3

Adrenaline mechanism "ABC of Adrenaline": Adrenaline--> activates Beta receptors--> increases Cyclic AMP

Carbon monoxide: electron transport chain target "CO blocks CO": Carbon monoxide (CO) blocks Cytochrome Oxidase (CO)

Metabolism sites "Use both arms to HUG": Heme synthesis Urea cycle Gluconeogenesis • These reactions occur in both cytoplasm and mitochondria.

Na/K pump: concentrations of Na vs. K on inside/outside of cell, pump action, number of molecules moved HIKIN': There is a HIgh K concentration INside the cell. From this can deduce that the Na/K pump pumps K into cell and Na out of cell. Alternatively: When I was learning this pump (circa 1992), a band that was "in" was Kris Kross, and a band that was "out" was "Sha Na Na Na": So pump moves K K (2 K) in and Na Na Na (3 Na) out. • Sadly, as infectious as their debut album was, Kris Kross can really no longer be classed as "in".

DNA bond strength (nucleotides) "Crazy Glue": Strongest bonds are between Cytosine and Guanine, strong like Crazy Glue (3 H-bonds), whereas the A=T only have 2 H-bonds. • This is relevant to DNA replication, as the weaker A=T will be the site where RNA primer makes the initial break.

Dicarboxylic acids (alpha, omega) C2 through C10: common names "Oh My, Such Good Apple Pie, Sweet As Sugar!": Oxalic Malonic Succinic Glutaric Adipic Pimelic Suberic Azelaic Sebacic

Infantile Beriberi symptoms Restlessness Sleeplessness Breathlessness Soundlessness (aphonia) Eatlessness (anorexia) Great heartedness (dilated heart) • Alternatively: Get 5 of 'em with BERI: Breathless/ Big hearted, Eatless, Restless, Insomnia.

AcetylCoA and acetacetylCoA are composed of "A Lighter Lease" (A LyTr LeIs): A=AcetylCoA or Acetoacetyl CoA Ly=Lysine Tr=Tryptophan Le=Leucine Is=Isoleucine

Fasting state: branched-chain amino acids used by skeletal muscles "Muscles LIVe fast": Leucine Isoleucine Valine

Glycolysis steps; "Goodness Gracious, Father Franklin Did Go By Picking Pumpkins (to) Prepare Pies": Glucose, Glucose-6-P, Fructose-6-P Fructose-1,6-diP, Dihydroxyacetone-P, Glyceraldehyde-P, 1,3-Biphosphoglycerate, 3-Phosphoglycerate, 2-Phosphoglycerate, (to), Phosphoenolpyruvate [PEP] Pyruvate; 'Did', 'By' and 'Pies' tell you the first part of those three: di-, bi-, and py-; 'PrEPare' tells location of PEP in the process.

Glycolysis Enzymes Mnemonic “Hungry Peter Pan And The Growling Pink Panther Eat Pies.” Hexokinase, Phosphohexo isomerase, Phosphofructokinase-1 (6-phosphofructo-1 kinase), Aldolase, Triose phosphate isomerase, Glyceraldehyde 3-phosphate dehydrogenase, Phosphoglycerate kinase, Phosphoglycerate mutase, Enolase, Pyruvate kinase

Citric acid cycle compounds "Cycles Involving Acids Suck Serious F@#%ing Monkey Organs": Citrate, Isocitrate, alpha-Ketogluterate, Succinyl CoA, Succinate, Fumerate, Malate, Oxaloacetate

Citric acid cycle compounds "Our City Is Kept Safe And Sound From Malice": Oxaloacetate, Citrate, Isocitrate, alpha-Ketoglutarate, Succinyl-CoA, Succinate, Fumarate, Malate

Citric acid cycle compounds "Can I Keep Selling Sex For Money, Officer?": Citrate, Isocitrate, alpha-Ketogluterate, Succinyl CoA, Succinate, Fumerate, Malate, Oxaloacetate

Citric acid cycle compounds "Oh! Can I Keep Some Succinate For Myself?": Oxaloacetate, Citrate, Isocitrate, alpha-Ketoglutarate, Succinyl CoA, Succinate, Fumarate, Malate

Citric acid cycle compounds "Oh Citric Acid Is Of (course) A SiLly STupid Funny Molecule": Oxaloacetate, Citrate, Aconitate, Isocitrate, Oxalosuccinate, Alpha-ketoglutarate, SuccinyL-CoA, SuccinaTe, Fumarate, Malate • SilLy and sTupid used to differentiate succinyL and succinaTe

Citric acid cycle compounds "Can I Ask Sharon Stone For My Orgasm?": Citrate, Isocitrate, Alpha-Ketoglutarate, Succinyl CoA, Succinate, Fumerate, Maleate, Oxaloacetate

Citric acid cycle compounds “Citrate is a key substrate for mitochondrial oxidation”: Citrate, Isocitrate, alpha-ketoglutarate, succinyl-CoA, Succinate, Fumarate, Malate, Oxaloacetate

Citrate Cycle Enzymes Mnemonic "Corrupt Anti Intelligence Agent Spoke Slander For Money." Citrate synthatase, aconitase, Isocitrate dehydrogenase, Alpha ketogluturate dehydrogenase, Succinyl CoA synthetase, Succinate dehydrogenase, Fumarase, Malate Dehydrogenase

Fabry's disease FABRY'S: Foam cells found in glomeruli and tubules/ Febrile episodes Alpha galactosidase A deficiency/ Angiokeratomas Burning pain in extremities/ BUN increased in serum/ Boys Renal failure YX genotype (male, X linked recessive) Sphingolipidoses

B vitamin names "The Rhythm Nearly Proved Contagious": • In increasing order: Thiamine (B1) Riboflavin (B2) Niacin (B3) Pyridoxine (B6) Cobalamin (B12)

Electron transport chain: Rotenone's site of action Rotenone is a site specific inhibitor of complex one.

Type 1 glycogen storage disease Type 1 = one (Von), ie Von Giereke's disease

Essential amino acids PVT. TIM HALL: Phe Val Thr Trp Ile Met His Arg Leu Lys • Pvt. is short for Private in the military. • Arg and His are considered semiessential. • Alternatively: "MATT VIL PHLy".

Golgi complex: functions "Golgi Distributes A SPAM": Distributes proteins and lipids from ER Add mannose onto specific lysosome proteins Sulfation of sugars and slected tyrosine Proteoglycan assembly Add O-oligosugars to serine and threonine Modify N-ologosugars on asparagine

Catabolism steps of branched chain amino acids "Truck hit the Ox to Death": Transamination Oxidative decarboxylation Dehydrogenation

Niacin deficiency: signs and symptoms The famous 4 D's: Diarrhoea Dermatitis Dementia Death (if untreated)

Enzymes: classification "Over The HILL": Oxidoreductases Transferases Hydrolases Isomerases Ligases Lyases • Enzymes get reaction over the hill.

Insulin: function INsulIN stimulates 2 things to go IN 2 cells: Potassium and Glucose.

Collagen concisely covered COLLAGEN: C-terminal propeptide (procollagen)/ Covalent Cross links/ C vitamin/ Connective tissue/ Cartilage/ Chondroblasts/ Copper Cofactor (Covalent Cross linking) Outside the cell is where collagen normally functions/ Osteoblasts/ Osteogenesis imperfecta Lysyl hydroxylase/ Lysyl oxidase (oxidatively deaminates lysyl and hydroxylysyl residues to form collagen cross links, last biosynthesis step) Long triple helical fibers/ Ligaments Alpha chains/ Attached by H bonds form triple helix/ Ascorbate for hydroxylation of lysyl and prolyl residues of pro-Alpha chains (postranslational modification) Gly in every third position/ Glycosylation of hydroxyl group of hydroxylysine with Glucose and Galactose; GOlgi allows procollagen to GO outside of cell Extracellular matrix/ Eye (cornea, sclera)/ Ehlers-Danlos Syndrome N-terminal propeptide (procollagen)/ Nonhelical terminal extensions • Note: Procollagen LEAVEs the cell to be cLEAVEd by procollagen peptidases

Essential amino acids "PVT. TIM HALL always argues, never tires": Phe Val Thr Trp Ile Met His Arg Lue Lys • Always argues: the A is for Arg, not Asp. • 'Never tires': T is not Tyr, but is both Thr and Trp.

Hypervitaminosis A: signs and symptoms "Increased Vitamin A makes you HARD": Headache/ Hepatomegaly Anorexia/ Alopecia Really painful bones Dry skin/ Drowsiness

Heme synthesis: amino acid precursors to basic unit of porphyrins, heme (pyrrole ring) "Dracula wants to Suck a Co-ed's blood [think heme] with his Glystening teeth!": Succinyl CoA and Glycine are precursor amino acids to pyrrole rings, which is the basic unit of porphyrins and heme.

Enzymes: competitive inhibitors "Competition is hard because we have to travel more kilometers (km) with the same velocity": With competitive inhibitors, velocity remains same but km increases

Phosphorylation cascade: action during low glucose "In the Phasted State, Phosphorylate": The phosphorylation cascade becomes active when blood glucose is low.

Sickle cell anemia: mutation "HbS isn't Very Good": At Sixth position of HB beta chain, Valine is present instead of Glutamic acid.

tRNA: molecular shape Fortunately, tRNA (Transfer RNA) is shaped like a capital T. • See diagram.

Malate-aspartate shuttle "MAD commute": Malate in. Alpha-ketoglutarate and D (Aspartate) out.

Vitamins: which are fat soluble KADE: Vitamin K Vitamin A Vitamin D Vitamin E

Phenylketonuria (PKU): which enzyme is deficient PHenylketonuria is caused by a deficiency of: Phenylalanine Hydroxylase

DNA/RNA/Protein

1. Molecular biology tools and techniques:

a.Cloning – introduction of pieces of DNA into a vector in order to permit amplification. Many methods of cloning exist. Commonly, total cellular DNA is cleaved, and each piece is inserted into a vector. The library of vectors is introduced into bacteria or another replication host. A bacteria which has a vector will then replicate, making many copies of the DNA in that vector, hence, a clone. (Lippincott page 404)

b.cDNA libraries – complementary DNA libraries are made by reverse transcribing (making DNA from RNA) all of the mRNA in a cell. The DNA copies are replicas of mRNA without introns. These can be used as probes, primers, or many other uses.

c.PCR-see page 146-biochemistry.

d.Restriction length fragment polymorphism—within the natural sequence of many genes are restriction sites, specific sequences cleaved by restriction enzymes. Many of these sites are polymorphic. That is, they contain differences which render them susceptible or perhaps not susceptible to cleavage at that particular site. These differences can be used to identify genes, or match DNA from different samples, as in forensics.

e.Sequencing – The major method of sequencing is the Sanger dideoxy nucleotide method. An elongation reaction is carried out using a primer just upstream of the portion to be sequenced. The mixture includes radioactive nucleotides except one of the nucleotides (A,T,G,or C) is dideoxy. That is, it does not have an oxygen at the 2 or 3 position of the ribose sugar. When one of the dideoxy bases is incorporated into the growing chain, elongation is stopped. A reaction is run containing dideoxy nucleotides of each base. For instance, one reaction contains all of the nucleotides but the adenosines are dideoxy. The results are then run on a gel. The sequence can be read by observing which bases were the terminating base at each position on the sequence.

2. Transcriptional regulation

a. The operon model – This model is related to the regulation of gene transcription (making RNA from genes) in certain environments. For instance the Lac operon consists of regulatory proteins that control production of proteins necessary to degrade lactose. These are only needed when lactose is present.

b. Eukaryotic transcription—Eukaryotic transcription is controlled by regions of DNA called promoters upstream from the material of the genes. Transcription factors bind to promoters, and help recruit RNA Polymerase II, which binds the TATA box, located approx. 25 bases upstream from the transcription start site. Another sequence, the CAAT box, is located approx. 40 bases upstream from the TATA box. Enhancer regions are other regions of DNA that bind specific protein that aid in transcription of certain gense. These regions can be located upstream, within the gene, within introns, close, or far from the transcription start site. Before a gene is transcribed a large complex of proteins is formed in the promoter region. The need for this large complex of proteins helps in gene regulation.

c. Role of steroid hormones – Steroid hormones cross membranes and travel directly to the nucleus of target cells. Bound to their respective receptors, steroid hormones act like transcription factors or enhancer binding proteins. They bind to hormone response elements near the genes they regulate and either enhance or inhibit transcription of those genes.

3. Translation (Protein synthesis)

a. Translation of mRNA occurs in the cytosol on ribosomes. Ribosomes can be free floating or attached to the ER membrane. Three nucleotides on mRNA encode for on amino acid. The start site is AUG on the RNA. This codes for methionine. Each amino acid is attached to a specific tRNA, which recognized the codon for that particular amino acid. Translation happens in three steps: initiation, elongation, and termination.

i.Initiation: Initiation involves assembling two ribosomal subunits, the mRNA, GTP, the tRNA with the first amino acid, and initiation factors that facilitate the whole process. The ribosome recognizes specific sequences on the mRNA and assembles the machinery. In bacteria, the first amino acid is N-formyl-methionine, while in eukaryotes it is normally methionine. The first tRNA with the appropriate amino acid is in the P-site of the ribosome, and the next tRNA with its appropriate amino acid arrives to the A-site. A peptide bond is formed between the two amino acids. Initiation factors aid in the setting up of the complex.

ii. Elongation: Elongation factors help the ribosome move down the mRNA with energy derived from GTP hydrolysis. tRNAs are attached to their amino acids using energy from ATP by specific synthetases for each amino acid and tRNA combination. Each time the ribosome moves down the mRNA, the nascent polypeptide is moved into the P-site, making room in the A-site for a new tRNA/amino acid pair.

iii. Termination: Termination occurs when the ribosome meets a termination sequence. Release factors cause the new peptide to be released from the ribosomes, and cause the dissociation of the ribosome complex.

b. Post translational modification of proteins occurs depending on the final destination and function. Modifications include trimming of proteins to active forms. Insulin for example, is synthesized as a zymogen and cleaved to the active molecule. Covalent alterations are also added to some proteins. These include glycosylation with different sugars, phosphorylation, hydroxylation, or association with coenzymes.

4. Acid-base titration curve of amino acid and proteins

a. Protons will dissociate from weak acids at a certain pH, depending on the strength of the bond of the dissociable hydrogen. This pH is called the pKa of the acid. The Henderson-Hasselbach equation relates the relative amount of acid and base at a given pH to the pKa of an acid.

b. Amino acids, since they have a carboxyl group, are weak acids. The pKa of most carboxyl groups is around 2, and the pKa for most amino groups of amino acids is around 9. They are referred to as pKa1 and pKa2 respectively. Some amino acids with an acidic or basic side chain have an additional pKa for the side chain hydrogen ion. See page 12 in Lippincott for examples of titration curves.

c. Proteins have titration curves as well. However, the carboxyl group and the amino group are the main titratable acids, as well as titratable side chains.

d. Titratable side chains include the acidic amino acids, aspartate and glutamate, basic amino acids arginine, lysine and histidine. The pKa of histidine is 6.0, so at physiologic pH it is not ionized.

5. Role of SH2 domains

a. The role of SH2 domains is simple: they bind phosphotyrosine. They are normally found in proteins involved in signal transduction. By binding to phosphotyrosine, they allow signals to be passed from one molecule to another. For instance, some receptors, when bound to a ligand, have tyrosine kinase activity. When these tyrosines are phosphorylated, and SH2 domain of another protein can bind to the cytoplasmic phosphotyrosine. The signal can be passed to other proteins and eventually to the nucleus.

Genetic Errors

1. Inherited hyperlipidemias-- Type Increased lipoprotein class Increased Lipid I Chylomicrons Triglycerides IIa LDL Cholesterol IIb LDL and VLDL Cholesterol and triglycerides III Remnants Triglycerides and cholesterol IV VLDL Triglycerides V VLDL and chylomicrons Triglycerides and cholesterol

2. Glycogen and lysosomal storage diseases are covered in First Aid.

3. Porphyrias --

a. Porphyrias are defects of porphyrin metabolism, leading to buildup of toxic metabolites. Porphyrins are ring structures. An example in heme without the iron. Many defects exist in many different steps in the pathways. They are classified depending on clinical and biochemical properties. Five main types exist:

i. congenital erythropoietic porphyria ii. erythrohepatic porphyria iii. acute intermittent porphyria iv. porphyria cutanea tarda v. mixed porphyria

b. Manifestations include light sensitivity, vesicles that heal when scarring, anemia. Pathogenesis is not well understood.

c. The clinical picture is someone with light sensitivity, so they only go out at night. They have weird fluorescing molecules in their bodies, so their teeth fluoresce (porphyrin rings.) Because of defects in heme synthesis and metabolism, anemia can be a problem, so they want to drink blood. Some say that vampire legends came from people with porphyrias. Interesting eh?

4. DNA repair defects (First Aid p. 149) Disease Features Type of repair defect Xeroderma pigmentosum (skin sensitivity to UV light) Skin tumors, photosensitivity, cataracts, neurological abnormalities Nucleotide excision repair defects, including mutations in helicase and endonuclease genes
Cockayne syndrome Reduced stature, skeletal abnormalities, optic atrophy, deafness, photosensitivity, mental retardation Defective repair of UV-induced damage in transcriptionally active DNA; considerable etiological and symptomatic overlap with xeroderma pigmentosum and trichothiodystrophy
Fanconi anemia Anemia; leukemia susceptibility; limb kidney, and heart malformations; chromosome instability As many as eight different genes may be involved, but their exact role in DNA repair is not yet known
Bloom’s syndrome (radiation)

• Growth deficiency, immunodeficiency, chromosome instability, increased cancer incidence Mutations in the reqQ helicase family

Werner syndrome Cataracts, osteoporosis, atherosclerosis, loss of skin elasticity, short stature, diabetes, increased cancer incidence; sometimes described as “premature aging” Mutations in the reqQ helicase family
Ataxia-telangiectasia (x-rays) Cerebellar ataxia, telangiectases*, immune deficiency, increased cancer incidence, chromosome instability Normal gene product is likely to involved in halting the cell cycle after DNA damage occurs
Hereditary nonpolyposis colorectal cancer Proximal bowel tumors, increased susceptibility to several other types of cancers Mutation in any of four DNA mismatch repair genes

• Telangectases are vascular lesions caused by the dilatation of small blood vessels. This typically produces discoloration of the skin.

Reference: Medical Genetics p. 39

5. Triplet repeat diseases. Disease Description Repeat sequence Normal & Abnormal range Parent in which expansion usually occurs Location of expansion Huntington disease Loss of motor control, dememtia, affective disorder CAG 6 to 34; 36 to >100 More often through father Exon Spinal and bulbar muscular atrophy Adult-onset motor-neuron disease associated with androgen insensitivity CAG 11 to 34; 40 to 62 More often through father Exon Spinocerebellar ataxia (type 1, 2, 3, 6) Progressive ataxia and other type specific symptoms CAG Varies with type More often through father Exon Dentatorubral-pallidoluysian atrophy/Haw River syndrome Cerebellar atrophy, ataxia, myoclonic epilepsy, choreoathetosis, dementia CAG 7 to 25; 49 to 88 More often through father Exon Myotonic dystrophy Muscle loss, cardiac arrhythmia, cataracts, frontal balding CTG 5 to 37; 100 to >1000 Either parent, but expansion to congenital form through mother 3’ Untrans-lated region Friedreich’s ataxia Progressive limb ataxia, dysarthria, hypertrophic cardiomyopathy, pyramidal weakness in legs GAA 7 to 22; 200 to 900 or more Disorder is autosomal recessive, to disease alleles are inherited from both parents Intron Fragile X syndrome (FRAXA) Mental retardation, large ears and jaws, macro-orchidism in males CGG 6 to 52; 200 to >2,000 Exclusively through mother 5’ Untras- lated region Fragile site FRAXE Mild mental retardation GCC 6 to 35; 200 or more More often through mother Unknown Reference: Medical Genetics p. 83

6. Inherited defects in amino acid metabolism. Name Prevalence Mutant gene product Chromosomal location Phenylketonuria (PKU) 1/10,000 Phenylalanine hydroxylase 12q24 Tyrosinemia (type 1) 1/100,000 Fumarylacetoacetate hydrolase 15q23-25 Maple syrup urine disease 1/180,000 Branched-chain -ketoacid decarboxylase (multiple subunits) Multiple loci Alkaptonuria 1/250,000 Homogentisic acid oxidase 3q2 Homocystinuria 1/340,000 Cystathionine -synthase 21q2 Oculocutaneous albinism 1/35,000 Tyrosinase 11q Cystinosis 1/100,000 Unknown 17p Cystinuria 1/7,000 SLC3A1 (type 1) 2p Reference: Medical Genetic p.138

Metabolism

1. Glycogen synthesis: regulation, inherited defects. A. Regulation: In the well fed state, Glycogen synthetase is allosterically activated by glucose 6-phosphate, as well as by ATP, a high energy signal in the cell. An elevated insulin level results in overall increased glycogen synthesis. Glucagon (in the liver) and Epinephrine (muscle and liver) bind cell membrane receptors and stimulate adenylate cyclase then cAMP. Glycogen synthetase is then phosphorylated by cAMP-dependent protein kinase which inhibits the production of glycogen. (Lippincott’s Biochem p. 142-145) B. Inherited defects: Glycogen storage diseases- First Aid p. 150

2. Oxygen consumption, carbon dioxide production, and ATP production for fats, proteins, and carbohydrates. A. Oxygen consumption occurs in the mitochondrial matrix. Cytochrome oxidase uses oxygen as the final electron acceptor and converts it to H2O. B. Carbon dioxide production results from reactions in several pathways including the TCA cycle and the Hexose Monophosphate Pathway (HMP). One molecule of CO2 (and one NADH) is produced in the conversion of Pyruvate to Acetyl-CoA by pyruvate dehydrogenase. Two molecules of CO2 are produced in the TCA cycle for every molecule of Acetyl-CoA. One molecule of CO2 (and one NADPH) is also produced by the conversion of 6-Phosphogluconate to Ribulose 5-phosphate in the HMP. C. ATP production Fats are broken down through Triacylglycerol degradation into fatty acids which are then broken down to Fatty Acyl CoA and then Acetyl-CoA which then enters the TCA cycle. Each Acetyl-CoA produces 3NADH, 1FADH2, 2CO2, 1GTP which is equal to 12ATP/acetyl CoA. Proteins are broken down to amino acids which enter the TCA cycle at different points. Refer to p.244 Fig. 22.22 in Lippincott’s Biochem for the metabolism of specific amino acids and associated genetic deficiencies. Carbohydrates are broken down to monosaccharides, the most common of which is D-glucose. Aerobic metabolism of glucose produces 38 ATP via malate shuttle, 36 ATP via G3P shuttle. Anaerobic glycolysis produces only 2 ATP per glucose molecule.

3. Amino acid degradation pathways (urea cycle, tricarboxylic acid cycle). Refer to Lippincott’s Biochem Figure 21.11, p.237 and Figure 22.2, p.244.

4. Effect of enzyme phosphorylation on metabolic pathways. Enzyme Enzyme Activity when Phosphorylated Description Glycogen phosphorylase Active Degrades glycogen Glycogen synthase Inactive Synthesizes glycogen Pyruvate kinase Inactive Converts Phosphoenolpyruvate (PEP) to Pyruvate Pyruvate dehydrogenase Inactive Converts Pyruvate to Acetyl-CoA Acetyl CoA carboxylase Inactive Converts Acetyl CoA to Malonyl CoA in triglyceride synthesis Hormone sensitive lipase Active Breaks triglyceride into a fatty acid and a diglyceride

Note: not a complete table - add enzymes as needed

5. Rate limiting enzymes in different metabolic pathways: First Aid p. 155

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

• Organ sites (First Aid 99 p 156):
• Liver: Most represented, including gluconeogenesis; fatty acid oxidation (-oxidation); ketogenesis;
• lipoprotein formation; urea, uric acid & bile acid formation; cholesterol synthesis.
• Brain: Glycolysis, amino acid formation.
• Heart: Aerobic pathways (e.g., -oxidation and Krebs cycle)
• Adipose tissue: Esterification of fatty acids and lipolysis
• Muscle: fast twitch: Glycolysis; slow twitch: Aerobic pathways
• Cell sites (First Aid 99 p 154):
• Mitochondria: -oxidation, acetyl-CoA production, Krebs cycle.
• Cytoplasm: Glycolysis, fatty acid synthesis, HMP shunt, protein synthesis (RER), steroid synthesis (SER).
• Both: Gluconeogenesis, urea cycle, and heme synthesis

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

• See liver diagrams for both states on page 159 of First Aid 99.
• Fed (Absorptive) state (BRS Biochemistry p 4):
• Glucose is oxidized by various tissues for energy or is stored as glycogen in liver and muscle. In liver, glucose is also converted to triacylglycerols, which are packaged in VLDL and released into the blood. Fatty acids of the VLDL and chylomicrons are stored in adipose tissue. Absorbed amino acids are used by various tissues to synthesis proteins, produce nitrogen-containing compounds, and produce energy.
• Fasting state (BRS Biochemistry p 7):
• With decreasing blood glucose level, the liver is stimulated by glucagon to supply glucose (glycogenolysis & gluconeogenesis) and ketones to the blood. The liver uses amino acids from muscles and fatty acids and glycerol from adipose tissue.
• Prolonged Fasting (BRS Biochemistry p 9):
• Muscles:  use of ketones &  oxidation of fatty acids for primary energy source.
• Brain: use of abundant ketones instead of glucose.
• Liver:  gluconeogenesis & spares muscle proteins.

8. Tyrosine kinases and their effects on metabolic pathways (insulin receptor, growth factor receptors)

• Insulin Receptor (Lippincott p 273):
• Insulin binding activates receptor tyrosine kinase activity in the intracellular domain of the -subunit.
• Tyrosine residues of the -subunit are autophosphorylated.
• Receptor tyrosine kinase phosphorylates other proteins, such as the insulin receptor substrate (IRS).
• Phosphorylated IRS promotes activation of other protein kinases and phosphatases, leading to the biological actions of insulin (see Topic 13 below).
• Insulin-like Growth Factor Receptor (BRS Physiology p 249):
• The IGF receptor has tyrosine kinase activity like the insulin receptor

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

• Considered “counterregulatory hormones” because they oppose many actions of insulin (Lippincott p 275):
• Glucagon: Acute, short-term regulation by stimulating hepatic glycogenolysis and gluconeogenesis.
• Epinephrine: Acute, short-term regulation by promoting glycogenolysis and lipolysis, inhibiting insulin secretion, and inhibits the insulin-mediated uptake of glucose by peripheral tissues.
• Cortisol: Long-term management by stimulating gluconeogenesis and lipolysis.
• Growth Hormone: Long-term management by stimulating gluconeogenesis and lipolysis.

10. Synthesis and metabolism of neurotransmitters.

• Acetylcholine (Correlative Neuroanatomy p 30):
• ACh is synthesized from acetyl-CoA and choline by the enzyme choline acetyltransferase in the presynaptic cholinergic nerve terminal.
• ACh is broken down after release into the synaptic cleft by the enzyme acetylcholinesterase.
• Choline is taken back up into the presynaptic nerve terminal to be converted back into ACh.
• Catecholamines (Correlative Neuroanatomy p 30):
• PhenylalaninetyrosineDOPAdopaminenorepinephrineepinephrine (First Aid 99 p151).
• Tyrosine is converted to DOPA by tyrosine hydroxylase.
• DOPA is converted to dopamine by DOPA decarboxylase.
• Dopamine is hydroxylated to NE and NE is converted to epinephrine by phenylethanolamine-N-methyltransferase.
• Dopamine and NE are inactivated by both MAO (presynaptic nerve terminal) and COMT (postsynaptic).
• Serotonin (Correlative Neuroanatomy p 32): Synthesized from the amino acid tryptophan.

11. Purine/pyrimidine degradation:

• Purine (G, A) degradation (BRS Biochemistry p 265):
• First phosphate and ribose are removed; then the nitrogenous base is oxidized.
• Guanine is degraded to xanthine and adenine to hypoxanthine, which is then oxidized to xanthine by xanthine oxidase (this enzyme requires molybdenum).
• Xanthine is oxidized to uric acid by xanthine oxidase.
• The kidneys excrete uric acid, which is not very water-soluble.
• Pyrimidine (C, U, T) degradation (BRS Biochemistry p 267):
• Unlike the purine rings, which are not cleaved in human cells, the pyrimidine ring can be opened and degraded to a highly soluble structures, such as -alanine and -aminoisobutyrate.
• The carbons produce CO2 and the nitrogens produce urea.

12. Carnitine shuttle: function and inherited defects.

• Function (Lippincott p 182): The carnitine shuttle transports the acyl group from cytosolic fatty acyl CoA molecules across the inner mitochondrial membrane, which is impermeable to CoA, returning it to mitochondrial CoA molecules. The newly formed mitochondrial fatty acyl CoA molecules can then undergo -oxidation.
• Inherited defects: The congenital absence of a carnitine acyltransferase in skeletal muscle, or low concentrations of carnitine due to defective synthesis, result in an inability to use long-chain fatty acids as a metabolic fuel, causing myoglobinemia and weakness following exercise.

13. Cellular/organ effects of insulin secretion (Lippincott p 273 and BRS Biochemistry p 154).

• Liver:  glycogen synthesis;  glucose production by inhibiting gluconeogenesis & glycogenolysis;  triacylglycerol synthesis & conversion to VLDL.
• Muscle:  glycogen synthesis;  glucose uptake by increasing the number of glucose transporters.
• Adipose tissue:  triacylglycerol degradation &  triacylglycerol synthesis;  glucose uptake by increasing the number of glucose transporters.
• Most tissues:  entry of amino acids into cells &  protein synthesis.
• Insulin does NOT significantly stimulate the transport of glucose into tissues such as liver, brain, & RBCs.

14. Effect of uncouplers on oxidative phosphorylation (Lippincott p 71).

• Compounds that increase the permeability of the inner mitochondrial membrane to protons can uncouple electron transport and phosphorylation.
• The energy produced by the transport of electrons, without a proton gradient, is released as heat rather than being used to synthesize ATP.
• 2,4-dinitrophenol, a lipophilic proton carrier that readily diffuses through the membrane, is an uncoupler.
• Aspirin in high doses (as well as other salicylates) is an uncoupler. This explains the fever that accompanies the toxic overdoses of these drugs.
• Uncoupling is different that just inhibiting electron transport, like cyanide does.

Biochemistry assays

Blots: function of Southern vs. Northern vs. Western "SN0W DR0P": • Match up the 1st word letter with 2nd word letter: Southern=DNA Northern=RNA Western=Protein • The 0's in snow drop are zeros, since there is no Eastern blot.