Exam 3: P450's and CBD

There is one b-type heme per polypeptide chain with an invariant cysteine as the axial ligand of heme iron

Smooth ER except for those involved in steroidogenesis. The microsomal P450s are bound to the ER membrane with their active site exposed to the cytosol. They are found in all tissues but most concentrated at entry points to the body (lungs, intestine, liver)

RH + O2+NADPH+ H+=>ROH + H2O + NADP+

The RH substrate binds to Fe3+. Oxygen binds Fe 3+ and then two electons are added to O2 to make superoxide and then the peroxide anion. This unstable anion allows the O2 molecule to be split leaving one O on the substrate and losing one in water.

NADPH=>NADPH cytochrome P450 reductase (FAD=> FMN)=> Cyt P450=> substrate. NADPH -450 reductase is an integral membrane protein that mediates electron transfer betweeen the two electron donor NADPH and the 1 electron acceptors

they penetrate protetive membranes and bind to proteins, reabosrb at the kidney, and accmulate in lipid soluble tissues. this is why detox invovles increasing the polarity of the xenobiotic

phase I= insertion of a polar group via P450s phase II=conjugation reactions to water soluble endogenous molecules to increase polarity and excretion

1. UDP-glucuronyl transferase adds glucuroinic acid 2. Glutathione S-transferase adds GS 3. Sulfotransferase asdds sulfate

Adds glucuronic acid to compunds with hydroxyl, amino, or sulfhydryl groups. The source of glucuronic acid is UDPGA

Catalyzes the nucelophilic attack of the thiolate anion of GSH on the electrophilic atom of any target lipophilic compound. Note that highly raactive epoxide intermediates are formed wihich have the potential to cause cell damage

Catalyzes the transfer of sulfate from PAPS to a nucelophilic acceptor, either a hydroxyl or amino group

The P450 oxygenates the xenobiotic into a conformationally hindered position that is inaccesible to the Phase II enzymes. The reactive intermediates can interact non-enzymatically with intracellular nucleophiles like proteins, DNA, and RNA to form covalent adducts resulting in injury and necrosis (examples: covalent adducts with guanosien teals to A G-C to T-A transversion, Benzo[a]pyrene is metabolized to the expoxide derivative a potent carcinogen)

Most common regulation is transcriptional with xenobiotic response elements functioning as transcription factors, may are subejct to developmental and tissue specific patterns of xpression

a measurable modification of the action of one drug by prior or concomitant admnistration of another drug, there is an increase in the level of one or more isoforms of the P450s in repsonse to the exposure to a drug

1. Mechaism based inhibtion 2. Disease states 3. Multiple drug therapy

Cetain drugs contain functional groups that are oxidized by P450s to metabolies that bind irreversibly to the enzyme resulting in inactivation of the enzyme

Disease with hepatic involvement such as cirhossis can impair drug elimination

Drugs compete fro the same P450, repression results in an increase in the half life of certain xenobiotics

When Ah ligands bind, the receptor complex enters the cell and binds to XREs that fucntion as transcriptional enchances in the nucelus. This results in production of mRNA for CYP1A1, the Ah receptor, and UDPglucuronyl trasferase. Note the TCDD is a very toxic chemical that induces CYP1A1 but is metabolized very slowly, TCDD (Dioxin) competes for normal ligands for the Ah receptor and inhibits normal functions

Acetaminophen (APAP) is metabolized by serveral pathwyas including glucoronidation, sulfation, and by CYP2E1. Normally only 5% of APAP is metabolized by CYP2E1 to NAPQI which is highly reactive. NAPQI is then rendered notoxic by congulation with GSH. IF there is an overdose of APAP, the first two pathways are exhauseted and moer NAPQI is generated. The GSH can't keep up and toxic NAPQI does damage to the liver cells.

treat with N-acetylcysteine (NAC) which is the precursor in synthesis of GSH. Enchanced production of GSH will allow more toxic NAPQI to be metabolised to APA conjugates.

In the fasting state when glucose is low, there are less precursors to make UDDPGA which is the necessary substrate for glucuronidation, the main conjugation pathwy for APAP detox. the APAP must be diverted to other pwathways, increasing the amount of NAPQI that is formed

0-12 hr non-specific Ab pain, vommiting, 12-24 hrs wose GI sxs, 24-48 hrs AST, ALT, maybe PT liver damage detectable >48 hrs altered mental state

Prothrombin/ clotting time. Fator 7 must be constatnly regenerated so it will be one of the first enzymes to indicate liver FUNCTION. AST and ALT will also indicate damage but give less info on fxn

therapeutic ratio=toxic dose/effective dose. A higher ratio is prefered because this indicates that the pt will have to take a large amount to reach toxicity compared to the small aount need for efficacy

greater than 4g/day for adults or 2.6 g/day for peds, 7-10 grams is a single toxic dose. Toxic therapeutic ratio is ~2

Two effects 1. Increases the CYP2E1 activity leading to more NAPQI productivity 2. IN chronics, low blood glucose means less precusors for UDPGA to make glucuronic acid

NAPQI damages the hepatocytes and stops the urea cycle. The build up of ammonia leads to the formation of glutamine in the brain which then leads to an increases osmotic pressure, brain swelling, and death

Grapefruit juice is rich in naringenin and quercitin which are effective inhibitors of CYP3A4 which metabolizes may perscription drugs (Statins, Calcium channel blocers) Ingestion of juice increases the levels of the drugs because the CYP3A4 is inhibited

They contain indoles and isothiocyantes whihc induce CYP1A1 in the intestines an aid in metabolism This decreased the exposure of interngal organs to dietary polyaromatic hydrocarbons

Iron deficiency leads to decreased activity of P450s resulting in increased elvels of drugs in the circulation

Riboflavin is converted to FMN and FAD which are the key electron carriers in cyt P450 reductase, part of the ETC needed for P450 function. In severe deficiencies, the activity of the P450s is decreased

The expression of P450s in liver is subejct to varous genetic polymorphisms. Certain induviduals/ ethnic groups cannot properly oxidize certain drugs because they lact the proper P450

7-14 drinks/week or >5 per ocasion can cause moderate fetal damage. Characteristics include growth deficiency, abnormal facies, mental retardation, attentional and behavioral problems

Ehtanol is absorbed in the stomach (10%) and the small intestine (90%) it is metabolized in the stomahc (10-20%) and liver (80-90%) via alcohol dehydrogenase and other enzymes

1. Alcohol dehydrogenase (liver)= Ethanol + NAD=> Acetaldehyde + NADH + H+ 2. Acetaldehyde dehydrogenase (mitochondria) Acetaldehyde + NAD+ + H2O=>acetate + NADH +H+

Acetaldehyde is very reactive, forms bonds with proteins impariing thier fxn, can damage liver leading to cell death

1. fatty liver 2. alcoholic hepatitis 3. Cirrhosis

alcohol and aldehyde dehydrogenase require NAD to break down ethanol. The enzymes of GNG and fatty acid oxidation also require NAD, the high concentration of NADH/ lack of NAD caused by ethanol inhibits GNG by preventing oxidation of lactate to pyruvate resuting in hypoglycemia and lactic acidosis

CHildren are much form dependend on GNG while fasting, Alcohol injgestion can cause sever hypoglycemia leading to irreversible brain damage

1. ketone bodies will form and are released into the blood enhancing the acidic condition 2. Processing of acetate is inhibited leading to the buildup of acetaldehyde which can cause cell damage

Vitamin A is converted to retinoic acid, an important signaling molecule, by the dame dehydrogenases that metabolize ethanol. Ethanol modulates specific P450sthat act on retinoic acid. Disruption in retionic acid signaling can contribute to fetal alcohol syndrome and development of many cancers