CHAPTER 10 Hemoglobin Metabolism

HEMOGLOBIN STRUCTURE
HEME BIOSYNTHESIS Hb first protein whose structure was described using x-ray crystallography conjugated globular 4 heme groups and 2 heterogeneous pairs of polypeptide chains 34 g/dL in RBC, 64,000 D transport oxygen from lungs to tissues modulates vascular dilation by transporting NO transports CO2 from tissues to lungs ********** HEME STRUCTURE protoporphyrin IX + Fe2+ = ferroprotoporphyrin 1 heme molecule = 1 oxygen molecule double bonds = red ********** GLOBIN STRUCTURE 4 globin chains in 1 heme consist of 2 identical pairs of unlike polypeptide chains, 141 to 146 amino acid each each chain is divided into 8 helices (AH) and 7 nonhelical segments ********** COMPLETE HB MOLECULE A. Primary structure amino acid sequence of polypeptide chains B. Secondary structure chain arrangements in helices and nonhelices C. Tertiary structure arrangement of helices in pretzel-like configuration D. Quaternary structure tetrameric molecule spherical, 4 heme groups attached to 4 polypeptide chains and may carry 4 molecules of oxygen 2 -globin and 2 non- globin chains occurs in mitochondria and cytoplasm of RBC bone marrow precursors (pronormoblast to reticulocyte) loss of mitochondria and TCA = no more Hb synthesis ********** GLOBIN BIOSYNTHESIS chromosome 16: and chromosome 11: , , , one copy of each globin gene per chromatid for a total of two genes per person w/ the exception of and (2 copies each per chromatid, total of4 genes per person) pronormoblast to reticulocytes arise via transcription of genetic code to mRNA and translation of mRNA to globin polypeptide chain in pronormoblasts: < translation: < when synthesized, chains are released from the ribosomes in the cytoplasm ********** HEMOGLOBIN ASSEMBLY HbA = 22 HbA2 = 2 2 HbF = 2 2 in F cells **********

HEMOGLOBIN ONTOGENY
chromosome 16: chromosome 11: , , first 3 months of embryonic development: and at birth: HbF is predominant

HbA1C most glycated Hb glucose attaches to N-terminal valine of the chain increased in diabetes mellitus **********

Each heme group is suspended between helices E and F Heme iron is positioned b/w 2 histidine radicals proximal bond F8 and distal histidine residue in E7 Distal histidine swings in and out to permit passage of oxygen Globin chain AAs in the cleft are hydrophobic. Outside AAs are hydrophilic (w/c makes it water soluble) **********

HEMOGLOBIN PRODUCTION REGULATION

HEME REGULATION key rate limiting step: Gly + succinyl CoA ALA heme inhibits a. transcription of ALAS gene b. ALA dehydrase c. porphobilinogen deaminase/hydroxymethylbilane synthase

HEMOGLOBIN BIOSYNTHESIS

 increased heme demand = increased ALAS synthesis Ferrochelatase heme synthase plays a regulatory role in heme biosynthesis inhibited by heme negative feedback or substrate inhibition by protoporphyrin IX ********** GLOBIN REGULATION Globin production is regulated by the rate at which the DNA is transcribed to mRNA globin=mRNAs Hemin Fe3+ oxidation product of heme impt in controlling the rate of globin synthesis in intact reticulocytes and various cell-free systems w/o it, inhibitor of globin synthesis accumulates Excess components of Hb (unpaired chains, protoporphyrin, iron) reduce RBC survival. ********** HEMOGLOBIN REGULATION Hb synthesis is stimulated by tissue hypoxia. During hypoxia, kidney produces increased EPO w/c stimulates Hb and RBC production. Reference intervals: Men: Women Newborns:

27mm Hg = 50% oxygen saturation of Hb shift to the left: <27 mmHg shift to the right: >27 mmHg 96% to 100% reference interval for arterial oxygen saturation shift to the left: higher % O2 saturation and affinity shift to the right: lower affinity Shift to the right lower affinity CO2, H+, Cl- (strengthen salt bridges) 2,3-BPG (R T) body temperature decreased pH presence of abnormal Hbs w/ low affinity for oxygen high fever acidosis hypoxia (high altitude) pulmonary insufficiency congestive heart failure severe anemia cardiac righ-to-left shunt Shift to the left body tempearature depleted 2,3-BPG alkalosis presence of Hb variants Hb: sigmoidal curve Mb: hyperbolic curve Myoglobin greater oxygen affinity than Hb 17000D monomeric releases oxygen only at very low partial pressures not as effective as Hb in releasing oxygen released when there is damage to the muscle like MI, trauma, severe muscle injury (rhabdomyolysis) excreted through the kidney levels may become elevated in renal failure HbF P50 of 19-21 mmHg left shift Hemoglobin also transport CO2 CO2 + H2O H2CO3 (carbonic anhydrase) H2CO3 H+ + HCO3(Cl shift, lungs) Carbaminohemoglobin lower affinity for oxygen than does Hb in the absence of CO2 **********

14-18g/dL (140-180 g/L) 12-15 g/dL (120-150g/L) 16.5-21.5 g/dL (165 to 215 g/L) **********

HEMOGLOBIN FUNCTION
Hb readily binds oxygen molecules in the lungs ( O2 affinity) and efficiently unload oxygen to the tissues (O2 affinity) 1.34 mL of oxygen per each gram of Hb Affinity of Hb depends on partial pressure of oxygen (PO2)

PO2 amount of oxygen needed to saturate 50% Hb P50 value Oxygen dissociation curve percent oxygen saturation of Hb versus PO2 Hb that is completely deoxygenated has little affinity for oxygen.. per oxygen molecule bound, avidity increases and HB then becomes fully oxygenated.

VARIANT HEMOGLOBINS
Bohr effect shifts in curve METHEMOGLOBIN

 separate different types of Hb ferric iron continuously being formed by spontaneous oxidation but fails to accumulate brownish to bluish shift to the left if >30% hypoxia and cyanosis increased: nitrites or metHb reductase activity, HbM disease assayed by spectral absorption analysis (CO-oximeter) peak: 620-640 nm at a pH of 7.1 methemoglobinemia may be treated by removal of offending substance or administration of ascorbic acid or methylene blue ********** SULFHEMOGLOBIN formed by irreversible oxidation of Hb by sulfonamides, phenacetin, acetanilide, phenazopyridine, created in vitro by addition of hydrogen sulfide to Hb greenish pigment if elevated cyanosis peak: 620nm does not shift when cyanide is added ********** CARBOXYHEMOGLOBIN combination of carbon monoxide w heme iron some are produced endogenously 541 nm cherry-red treatment: hyperbaric oxygen

Carbon monoxide silent killer odorless and colorless affinity is 240x, release is 10,000x slower exposure may be coincidental, accidental or intentional (suicidal) toxic effects: headaches, dizziness, coma, convulsions **********

HEMOGLOBIN MEASUREMENT
CyanmetHb method 1. Lysing agent frees Hb 2. Free Hb combines w/ potassium ferricyanide contained in cyanmetHb reagent 3. Ferrous to ferric metHb 4. MetHb + potassium cyanide cyanmetHb 5. CyanmetHb measured at 540nm

Sodium lauryl sulfate (SLS) convert Hb to SLS-metHb does not generate toxic wastes Hb electrophoresis