Subhadipa 2020 Structure of Hemoglobin What is Hemoglobin? Subhadipa 2020 • Hemoglobin is a two-way respiratory carrier, transporting oxygen (as oxyhemoglobin) from the lungs to the tissues and facilitating the return transport of carbon dioxide (as carbamino hemoglobin). • In the arterial circulation, hemoglobin has a high affinity for oxygen and a low affinity for carbon dioxide , organic phosphates, and hydrogen and chloride ions. • Normal concentration of hemoglobin (male)—14-16 gm % and Normal concentration of hemoglobin (female)—13-15 gm %. • Normal hemoglobin means 97% HbA +2% HbA2 + 1% HbF. • Glycated Hemoglobin or HbA1C (α 2β2) is found > 5% in diabetes mellitus patients. • It is composed of 4 globin chains and porphyrin ring with central iron. • Iron must be in ferrous state which can binds to oxygen and form oxy hemoglobin. Ferric state can’t carry oxygen and its form methemoglobin. Structure of Hemoglobin Subhadipa 2020 • Hemoglobin comprises four subunits, each having one polypeptide chain and one heme group. Each subunit has a molecular weight of about 16,000 daltons , for a total molecular weight of the tetramer of about 64,000 daltons. • Haemoglobin, the red pigment in blood, consists of a protein component and the iron complex of a porphyrin derivative : Haemoglobin = globin (protein) + haemochromogen (Fe (II) complex) • There are four heme group each attached to one globin chain . So, one Hb molecule can carry 4 oxygen molecules. • All hemoglobins carry the same prosthetic heme group iron protoporphyrin IX (4%) associated with globin (96%) made up of polypeptide chain of 141 (alpha) and 146 (beta) amino acid residues. • The ferrous ion of the heme is linked to the imidazole N of two histidine residues of the polypeptide chain. • The porphyrin ring is wedged into its pocket by a phenylalanine of its polypeptide chain. Structure of Hemoglobin Subhadipa 2020 A. Globin Chains • The polypeptide chains of adult hemoglobin themselves are of two kinds, known as alpha and beta chains , similar in length but differing in amino acid sequence. • The alpha chain of all human hemoglobins, embryonic and adult, is the same . • The non-alpha chains include the beta chain of normal adult hemoglobin (HbA; α2β2), the gamma chain of fetal hemoglobin (HbF; α2γ2), and the delta chain of minor adult hemoglobin (HbA 2; α2δ2). • Sickle cell hemoglobin is denoted as α2S2 (HbS). • In alpha chain, there are 38 histidine residues (has buffering action). 58 th is distal and 87 th is proximal histidine in respect of ferrous ion. Globin gene clusters Subhadipa 2020 Normal hemoglobin variants and subunits Subhadipa 2020 Structure of Hemoglobin Subhadipa 2020 B. Protoporphyrin IX ring • Prosthetic group consists of ferrous ion and protoporphyrin IX ring (cyclic). • Protophorphyrin consists of tetrapyrroles. • Tetrapyrrole are linked by methane (=CH-H) bridges producing a conjugated double bond system. • Ferrous ion linked to 4 N of pyrroles. • Pyrrole I (A) and II (B) have the methyl and vinyl side chains. • Pyrrole III (C) and IV (D) have the methyl and propionate side chains. • Heme is the site of reversible O2 attachment . Four levels structure of Hemoglobin Subhadipa 2020 • Primary Structure At its simplest level, hemoglobin is made up of amino acids in chains. These chains are polypeptides that are also stuck to a heme molecule, which is where the oxygen will eventually stick. Hemoglobin is different than other proteins because its individual polypeptides, of which there are four, are called globins instead of simply protein subunits. • Secondary Structure The 7 alpha helices and short non helical randomly coiled segments. • Tertiary Structure The heme molecule is important for the tertiary bending structure of hemoglobin, as it helps twist the globins into shape by connecting to histidine residues on them. Non polar residues are located deep inside the coils, polar residues are at the outside. The heme entity is buried into the hydrophobic pocket. • Quaternary Structure Of the four globins that make up hemoglobin, two are identical and called alpha chains, and the other two are called beta chains and are also identical. They can also be called alpha- globins and beta-globins. The four peptides packed tightly together in a tetrahedral array to form an overall spherically shaped structure that is held together by hydrophobic interaction, some hydrogen bonds and salt bridges. Subhadipa 2020 R and T state of Hemoglobin • There are two states in the hemoglobin, the T state (the tense state) and the R state (the relaxed state). • The T state has less of an affinity for oxygen than the R state . In the concerted mode of cooperativity, the hemoglobin must either be in its T state or R state. • Salt bridges (thin lines) linking the subunits in the T structure break progressively as oxygen is added , and even those salt bridges that have not yet ruptured are progressively weakened (wavy lines). • The transition from T to R does not take place after a fixed number of oxygen molecules have been bound but becomes more probable as each successive oxygen binds. • The transition between the two structures is influenced by protons, carbon dioxide, chloride, and BPG; the higher their concentration, the more oxygen must be bound to trigger the transition. • Fully oxygenated molecules in the T structure and fully deoxygenated molecules in the R structure are not shown because they are unstable. Sickle cell hemoglobin Subhadipa 2020 • Sickle hemoglobin differs from normal hemoglobin by a single amino acid: valine replaces glutamate at position 6 on the surface of the beta chain. • Sickle cell anemia is a blood disease that affects red blood cells. Normal red blood cells are round. In people with sickle cell anemia, hemoglobin – a substance in red blood cells – becomes defective and causes the red blood cells to change shape. The faulty hemoglobin is called hemoglobin S (HbS), and it replaces normal hemoglobin which is called hemoglobin A (HbA). Over time, the red blood cells become rigid and shaped like crescent moons or sickles. • The sickle-shaped red blood cells: i. Clog blood vessels, causing episodes of pain and cutting off oxygen to tissues and organs. ii. Get trapped in the spleen (an organ that gets rid of old cells) where they are destroyed. The body cannot replace the lost cells fast enough. As a result, the body has too few red blood cells, a condition known as anemia..