Studies of the oxidation states of hemoglobin M Boston and hemoglobin M Saskatoon in blood by EPR spectroscopy.

The extent of the oxidation of Hemoglobin (Hb) M Saskatoon (beta 63His-->Tyr) and Hb M Boston (alpha 58His-->Tyr) in the patient's blood was determined by measurement of the intensity of EPR signals at g perpendicular = 6.0 for the normal subunits, g1 = 6.7 for the mutant subunits of Hb M Saskatoon and g1 = 6.3 for those of Hb M Boston, respectively. The amounts of reduced mutant subunits were estimated from the EPR signal intensities and the amounts of Hb present as mutant Hb in the blood. About 50% and 76% of mutant subunits in Hb M Boston and Hb M Saskatoon remained reduced in the fresh blood. Gentle shaking of the blood at 37 degrees C for 15 hours in air brought about autoxidation of the normal subunits as well as the mutant subunits of the two Hbs M, indicating that the presence of the mutant subunits facilitated autoxidation of the normal subunits. Possible involvement of NADH-metHb reductase in erythrocytes in maintenance of the reduced mutant subunits of Hb M Saskatoon was discussed.

Unusual CO bonding geometry in abnormal subunits of hemoglobin M Boston and hemoglobin M Saskatoon.

To clarify the role of the proximal histidine (F8-His), distal His (E7-His), and E11 valine (E11-Val) in ligand binding of hemoglobin (Hb), we have investigated the resonance Raman (RR) spectra of the carbon monoxide adduct of Hbs M (COHb M) in which one of these residues was genetically replaced by another amino acid in either the alpha or beta subunit. In the fully reduced state, all Hbs M gave v3 at approximately 1472 cm-1 and vFe-His at 214-218 cm-1, indicating that they have a pentacoordinate heme and the heme iron is bound to either E7-His or F8-His. The porphyrin skeletal vibrations of the COHb M were essentially unaltered by replacements of E7- or F8-His with tyrosine (Tyr) and of E11-Val by glutamic acid (Glu). The vCO, vFe-CO, and delta Fe-C-O frequencies of COHb M Iwate (alpha F8-His----Tyr), COHb M Hyde Park (beta F8-His----Tyr), and COHb M Milwaukee (beta E11-Val----Glu) were nearly identical with those of COHb A. In contrast, the RR spectra of COHb M Boston (alpha E7-His----Tyr) and COHb M Saskatoon (beta E7-His----Tyr) gave two new Raman bands derived from the abnormal subunits, vFe-CO at 490 cm-1 and vCO at 1972 cm-1, in addition to those from the normal subunits at 505 cm-1 (vFe-CO) and 1952 cm-1 (vCO). The CO adduct of the abnormal subunits exhibited apparently no photodissociation upon illumination of CW laser with a stationary cell under which the normal subunit exhibited complete photodissociation.(ABSTRACT TRUNCATED AT 250 WORDS)

Enhanced reaction with Vicia graminea lectin and exposed terminal N-acetyl-D-glucosaminyl residues on a sample of human red cells with Hb M-Hyde Park.

A sample of polyagglutinable red cells was obtained from a healthy individual (group O, N) possessing a hemoglobin (Hb) variant called Hb M-Hyde Park. The sialic acid content of the individual's red cells is 90 percent of normal, and his cells are agglutinated by monoclonal but not lectin anti-Tn, a panel of lectins specific for N-acetylgalactosamine (or galactose), and N-acetylglucosamine. Enhanced agglutination reactions were obtained with Vicia graminea, Ulex europaeus, and human anti-I and -i. Using various enzyme treatments and different methods of labeling cell surface components, two defective cell membrane sites have been identified: one associated with the O-linked oligosaccharides on sialoglycoproteins and the other associated with exposed N-acetylglucosaminyl residues located on membrane components of apparent molecular weights 88,000 to 130,000 and 46,000 to 73,000 (probably the Band 3 and Band 4.5 regions, respectively).

Hb M Milwaukee in a German family.

The second occurrence of Hb M Milwaukee is reported in two members of a German family who had cyanosis since early childhood. Contrary to earlier reports, Hb M Milwaukee exhibits a distinct heat instability. It is suggested, that in this family the variant resulted from a new mutation.