Biochemistry_of_Blood_I_FA23_LEO (1).pptx

Biochemistry of Blood I: Heme Synthesis VA N E S S A D E L A R O S A , P H D CV I FA 2 0 2 3 Session Objectives •Describe heme synthesis, regulation of the rate-limiting step catalyzed by 5-aminolevulinic acid (δALA) synthase (ALAS), connections to the TCA cycle and amino acid metabolism, and the...

Biochemistry of Blood I: Heme Synthesis VA N E S S A D E L A R O S A , P H D CV I FA 2 0 2 3 Session Objectives •Describe heme synthesis, regulation of the rate-limiting step catalyzed by 5-aminolevulinic acid (δALA) synthase (ALAS), connections to the TCA cycle and amino acid metabolism, and the requirement for iron. •Discuss how lead poisoning affects heme biosynthesis and mimics porphyria. •Explain heme structure and function, primary sites of synthesis and catabolism, and associated clinical disorders. •Identify porphyrias resulting from enzyme deficiencies in heme synthesis, such as Acute Intermittent Porphyria (AIP), Porphyria Cutanea Tarda (PCT), and Erythropoietic Protoporphyria (EPP) •Predict accumulating upstream intermediates in porphyrias that can be cytotoxic in the presence of light and molecular oxygen. •Outline the relationship between X-linked Sideroblastic anemia and ALAS2. Heme overview •Heme metabolism controlled at multiple levels •Heme is synthesized in mitochondria •Heme is derived from metabolic intermediates •Heme required for many oxygen binding proteins and enzymes • hemoglobin, myoglobin, cytochrome P450 enzymes •Heme degradation produces bilirubin •Porphyrias are inherited & acquired disorders of heme synthesis Porphyrins Cyclic tetrapyrroles capable of chelating various metals to form essential prosthetic groups for various biological molecules Heme consists of a protoporphyrin IX ring chelated with Fe2+ Heme is the most common porphyrin found in the body Heme can rapidly autooxidize to hemin (Fe3+) Heme synthesis overview • Pathway partially regulated by compartmentalization of different steps (mito vs. cyto) • 8 enzymatic steps • ALAS is rate limiting step • Excess heme is TOXIC 5-aminolevulinate synthase (ALAS) Glycine & succinyl-CoA (TCA cycle intermediate) precursors Requires PLP (vitamin B6) cofactor Regulated in part by differential expression of housekeeping (ALAS1) & erythroid-specific (ALAS2) isozymes ALAS1 expressed in liver/other tissues ALAS2 expressed in erythroid cells ALAS1 regulated by heme Liver is major site of heme synthesis for heme proteins ALAS1 is feedback-regulated by the intracellular concentration of heme. 1. Hemin inhibits transcription of ALAS1 2. Free heme inhibits translation of ALAS1 3. Free heme inhibits translocation of ALAS1 enzyme into mitochondria ALAS2 regulated by iron •ALA‐S2 is regulated post‐ transcriptionally by cellular iron status •ALAS2 mRNA has iron response element (IRE) •IRP (iron binding protein) can bind the IRE to inhibit translation of genes •Heme and globin synthesis is highly coordinated in erythroid cells High intracellular iron favors ALAS2 mRNA translation Heme synthesis can proceed and is available for normal hemoglobin production Low intracellular iron inhibits ALAS2 mRNA translation Heme is not available for hemoglobin synthesis ALA dehydratase (ALAD) •Condenses two molecules of ALA to form porphobilinogen (PBG) •Cytosolic enzyme containing Zn2+ •Lead & other heavy metals can inhibit enzyme by displacing Zn2+ •Inhibition responsible for the elevation in ALA and the anemia seen in lead poisoning Porphyrin formation • Enhances rate of Fe2+ insertion into protoporphyrin IX • Regulated by free heme • Inhibited by lead PPO converts protoporphyrinogen IX to protoporphyrin IX CPOX located in mitochondrial intermembrane space converts propionic acid side chains (P) to vinyl groups (V) to form protoporphyrinogen IX UROS directs stereochemistry of reaction to form uroporphyrinogen III porphobilinogen deaminase (PBGD; uroporphyrinogen I synthase) catalyzes head-to-tail condensation of four porphobilinogens UROD decarboxylates uroporphyrinogen III to form coprophorphyrino gen III (& I) Porphyrias •Deficiencies or mutations of enzymes in the synthetic pathway of heme lead to the types of porphyria •Overaccumulation of porphyrin •XLP is an exception caused by gain of function mutation Porphyrias •Classified depending on target tissue where the enzyme deficiency occurs •Classified as chronic or acute •Clinical manifestations result from accumulation of porphyrins or their precursors Z. Karim et al. 2015 Photosensitivity •Protoporphyrin abnormally accumulates in certain tissues of the body, especially the blood, liver, and skin. •Protoporphyrin molecules absorb energy from sunlight (photoactivation)  damage to the skin •UV irradiation of porphyrins in presence of molecular oxygen generates dangerous ROS (singlet oxygen) that can produce cellular damage  photosensitivity •Specific to porphyrias that accumulate intermediates AFTER tetrapyrrole ring closure Brown et al., 2018 Hepatic porphyrias • Increased synthesis of intermediates that occur prior to the genetic block • ⇑ ALAS1 activity (due to ⇓ regulatory heme) = ⇑ accumulation of pathway intermediates • Porphyria cutanea tarda is most common porphyria Hepatic porphyrias ACUTE INTERMITTENT PORPHYRIA (AIP) PORPHYRIA CUTANEA TARDA •Acute •Alcohol •Lack of CYP450 activity •Hepatitis C infection •Triggered •Photosensitivity •Neuropathy •Dark red urine •Lack of photosensitivity Erythroid porphyrias • Heme synthesis in erythroid cells of the bone marrow affected • ⇑ porphyrins (bone marrow, RBCs, plasma, urine, feces, teeth, bone) • ⇑ ALAS2 activity due to ⇑ iron (inactive IRP) • Can lead to iron overload XLPP • Hepatic porphyria • ALAS2 gain of function Overproduction of ALAS2 • Photosensitivity • No iron overload XLSA • NOT a porphyria, but results in microcytic anemia that is vitamin B6-responsive • ALAS2 loss-of-function • Iron overload= reduced heme synthesis stimulates erythropoiesis, increasing iron turnover and causing non-ferritin iron accumulation • “ring” around nucleus due to iron-laden mitochondria Lead poisoning •ALAD & ferrochelatase inhibited by lead •Displaces zinc as co-factor in ALAD & inhibits ferrochelatase •Manifests with features of porphyrias •ALA and/or protoporphyrin accumulate in urine •Treatment: lead chelation Summary