Charlotte Head

A Doubly Cross-linked Human Hemoglobin EFFECTS OF CROSS-LINKS BETWEEN DIFFERENT SUBUNITS

Human deoxyhemoglobin cross-linked with trimesyl tris(3,5-dibromosalicylate) produces the previously reported cross-linked hemoglobin in which the ϵ amino groups of the two β chain 82 lysyl residues are joined by a trimesyl bridge. Further specific modification of this protein directed to the α subunits with bis(3,5-dibromosalicyl)fumarate gives a doubly cross-linked material in which the ϵ-amino groups of the two α chain 99 lysyl residues are now joined by a fumaryl bridge. The singly cross-linked β chain species binds oxygen cooperatively with a high oxygen affinity (P50 = 4.8 torr at pH 7.4). The addition of the second cross-linking reduces the oxygen affinity to 15.9 torr, which compares with 13.0 torr for the singly cross-linked α chain species. The doubly cross-linked hemoglobin retains significant cooperativity with a Hill coefficient of 2.3 compared with 3.0 for unmodified hemoglobin. Because some of the groups responsible for the Bohr effect are acylated, this doubly cross-linked hemoglobin exhibits 25% of the normal Bohr effect and less than 20% of the normal chloride effect. The use of two distinct cross-links within the same tetramer provides a material for physical and structural analysis as well as for further modifications for specific applications. The results indicate that the cross-link introducing the lowest oxygen affinity in the two singly cross-linked species appears to control the overall affinity in this doubly cross-linked species.

HB Washtenaw [βT11(A8)VAL-PHE]: An Electrophoretically Silent, Unstable, Low Oxygen Affinity Variant Associated with Anemia and Chronic Cyanosis

Hb Washtenaw [beta 11(A8)Val-->Phe] is a new, low oxygen affinity variant with a previously undescribed substitution, identified in seven members over three generations of a Hungarian-American family. The hemoglobin is mildly unstable and the family members studied are clinically asymptomatic but mildly cyanotic, and some exhibit mild anemia. The index case had severe pulmonary hypertension and some of the family members had increased pulmonary vascular resistance on echocardiography. An association between the inheritance of this abnormal hemoglobin and the pathogenesis of primary pulmonary hypertension is suggested but the mechanism is unclear.

A New Electrophoretically Silent Variant That Causes Erythrocytosis

A high oxygen affinity hemoglobin, previously undescribed, was found in a healthy, asymptomatic patient with mild erythrocytosis and left-shifted hemoglobin-O2 dissociation curve. The hemoglobin variant could not be distinguished from Hb A by any of several electrophoretic methods nor by ion exchange chromatography. It was separated and analyzed by reversed phase high performance liquid chromatography. Structural analysis revealed the substitution beta 109 (G11) Val----Leu. The variant was named Hb Johnstown. The amino acid substitution perhaps disrupts alpha 1 beta 1 contacts in the deoxyhemoglobin conformation, thus shifting the equilibrium towards the high affinity oxyhemoglobin conformation.

HB Seal Rock [(α2)142 Term→glu, Codon 142 TAA→GAA]: An Extended α Chain Variant Associated with Anemia, Microcytosis, and α-Thalassemia-2 (-3.7 KB)

Hb Seal Rock was first reported in a young African-American woman and her 2-year-old daughter (1). It is an extended α chain variant which, like Hb Constant Spring, is present in small quantity and is expressed as an α thalassemia. The mutation, TAA→GAA affects codon 142 of the α2 gene. In this family, the index case was a compound heterozygote for Hb Seal Rock trait and for α-thalassemia trait(-3.7 kb).Her hematologic expression was similar to mild Hb H disease, presumably because the Seal Rock mutation affects the α2 gene that is normally responsible for approximately 70% of α-globin synthesis. Her daughter had only Hb Seal Rock trait, but was phenotypically α-thalassemia-2 trait due to the expression of the Seal Rock mutation on one of her α2-globin genes, the other three α-globin genes being unaffected.

Hemoglobin Linkoping [β36 (C2) Pro→Thr] in a Large Finnish Family from Astoria, Oregon, USA

Eleven members of a large Finnish family from Astoria, Oregon were studied because of an erythrocytosis. No abnormality was detected by the usual hemoglobin electrophoretic tests, but an abnormal variant was separated by reverse phase HPLC. All of the affected individuals have an increased oxygen affinity with a P50 for whole blood at 37 degrees C averaging 18 torr. Fifty percent of their hemoglobin was found to have a threonyl residue in place of the normal prolyl residue at position 36 (C2) of the beta globin chain. This abnormality is identical to Hb Linkoping which was recently reported in a Finnish man living in Sweden.

Hemoglobin Denver [α2β241 (C7) Phe→Ser]: A Low-O2-Affinity Variant Associated With Chronic Cyanosis and Anemia

Objective To report a previously undescribed low-O 2 -affinity hemoglobin variant that is associated with chronic cyanosis. Design Pertinent laboratory and historical data for the index case (from Denver, Colorado) and certain family members were recorded, and the hemoglobin variant was characterized. Material and Methods Electrophoresis, high-performance liquid chromatography (HPLC), and isoelectric focusing were used to examine blood specimens for the presence of hemoglobin Variants, and the O 2 affinity of whole blood was determined. The abnormal peptide detected on reverse-phase HPLC of separated globin chains was analyzed for its amino acid composition and sequence. Results Although no abnormal hemoglobin band separated from hemoglobin A on electrophoresis, HPLC, and isoelectric focusing, a heat test showed hemoglobin instability, and O 2 affinity studies disclosed an appreciably right-shifted dissociation curve. On chromatography, the new variant—hemoglobin Denver—was found to be due to a substitution of serine for phenylalanine at position 41 (C7) in the β chain. In addition to substantial reduction in O 2 affinity, hemoglobin Denver is accompanied by moderate reticulocytosis and mild anemia. Conclusion Hemoglobin Denver causes no clinical symptoms other than cyanosis, which is attributable to the low O 2 affinity.

Hemoglobin Columbia Missouri or α2[88 (F9) Ala→Val]β2: A New High-Oxygen-Affinity Hemoglobin That Causes Erythrocytosis

A previously undescribed hemoglobin variant, hemoglobin Columbia Missouri, α88 (F9) Ala→Val, was detected in a 22-year-old white man who was undergoing assessment for erythrocytosis. This new hemoglobin variant does not separate from hemoglobin A by electrophoresis in conventional media, by isoelectric focusing, or by electrophoresis of purified globin chains in 8 M urea. It exhibits a high oxygen affinity, with a P50 (oxygen tension at 50% saturation) of 19.3 torr for the patients whole blood. The substitution of hemoglobin Columbia Missouri is an internal residue near the end of the F helix of the α chain. Hemoglobin Okazaki has an arginyl residue substitution for a cysteinyl residue at F9 (β93) in the β chain. In comparison with hemoglobin Okazaki, the substitution in hemoglobin Columbia Missouri has a more pronounced effect on oxygen affinity. Consequently, hemoglobin Columbia Missouri is associated with erythrocytosis, whereas hemoglobin Okazaki is not.

Two Families With Hemoglobin Sogn, β(A11)14 Leu→Arg, in Minnesota and Indiana: Hematologic, Functional, and Biosynthetic Features

Sogn hemoglobinopathy was identified in a young American woman and in a young American man of apparently unrelated families of Norwegian ancestry. Both persons were asymptomatic and without clinical or hematologic manifestations. Hemoglobin Sogn, beta(A11)14 Leu----Arg, is an unstable hemoglobin that may easily be mistaken for hemoglobin S, G, or D by alkaline hemoglobin electrophoresis. These are the first known instances of hemoglobin Sogn outside of Norway. Oxygen affinity is normal. Sogn hemoglobinopathy is an incidental finding that has no adverse implication for the health of heterozygotes.