Hemoglobin stability

Hematological and Hemoglobin Stability Data on 22 Members of the Family with Hb-Atlanta... 

Ligler FS, Stratton, LP, Rudolph AS. Liposome encapsulated hemoglobin stabilization, encapsulation, and storage. Prog Clin Biol Res 1989 319 435. 

Binding of protons to critical groups on hemoglobin stabilizes deoxy-bemoglobin and thereby decreases the O2 binding affinity of hemoglobin. 

R17. Rieder, R. E., Hemoglobin stability, observations on the denaturation of normal and abnormal hemoglobins by oxidant dyes, heat, and alkali. J. Clin. Invest. 49, 2369-2376 (1970). 

The allosteric effect is seen in hemoglobin which can exist in two quaternary stmctural states oxygenated (R) or deoxygenated (T). The binding of one O2 or some other effector to one of the subunits stabilizes the R form as compared to the T form. Binding of a second and third O2 stabilizes it even further. 

Reactivity. Hemoglobin can exist ia either of two stmctural coaformatioas, corresponding to the oxy (R, relaxed) or deoxy (T, tense) states. The key differences between these two stmctures are that the constrained T state has a much lower oxygen affinity than the R state and the T state has a lower tendency to dissociate into subunits that can be filtered in the kidneys. Therefore, stabilization of the T conformation would be expected to solve both the oxygen affinity and renal excretion problems. 

The transition between the T and R states of hemoglobin is also deeply involved in the Bohr effect and cooperativity. Therefore stabilization of either of the two stmctures should diminish these effects, which have important physiologic consequences. The clinical consequences of stabilization are not known. 

An interesting property of DBBF—hemoglobin is its thermal stability. This property has been used to achieve both a partial purification of the cmde reaction mixture after cross-linking and inactivation of vimses in the final product (101,102). 

This reaction is driven to the right in tissues by the high COg concentration the equilibrium shifts the other way in the lungs where [COg] is low. Thus, car-bamylation of the N-termini converts them to anionic functions, which then form salt links with the cationic side chains of Arg al41 that stabilize the deoxy or T state of hemoglobin. 

Residue H21 of the y subunit of fetal hemoglobin (HbF) is Ser rather than His. Since Ser cannot form a salt bridge, BPG binds more weakly to HbF than to HbA. The lower stabilization afforded to the T state by BPG accounts for HbF having a higher affinity for Oj than HbA. 

In hemoglobin M, histidine F8 (His F8) has been replaced by tyrosine. The iron of HbM forms a tight ionic complex with the phenolate anion of tyrosine that stabilizes the Fc3 form. In a-chain hemoglobin M variants, the R-T equilibrium favors the T state. Oxygen affinity is reduced, and the Bohr effect is absent. P Ghain hemoglobin M variants exhibit R-T switching, and the Bohr effect is therefore present. 

Other tests useful for the detection of unstable hemoglobins are the heat stability test, the Isopropanol precipitation test, and the PCMB or PMB precipitation test ... 

The stability of hemoglobin In a freshly prepared red cell hemolysate at temperatures varying between 50 and 70 Is distinctly less than that of a normal control. The unstable variant will also show decreased stability at 37 In the presence of the Isopropanol-trls buffer, pH 7.4. 

Closely monitor patients for efficacy and toxicity while they are receiving hydroxyurea. Monitor mean corpuscular volume (MCV) because it increases as the level of HbF increases. If the MCV does not increase with hydroxyurea use, the marrow may be unable to respond, the dose may not be adequate, or the patient may be noncompliant.27 HbF levels also can be monitored to assess response. Assess blood counts every 2 weeks during dose titration and then every 4 to 6 weeks once the dose is stabilized. Temporary discontinuation of therapy is warranted if the hemoglobin level is less than 5 g/dL (50 g/L or 3.1 mmol/L), the absolute neutrophil count is less than 2000/mm3 (2 x 109/L), platelets are less than 80,000/mm3 (80 x 109/L), or reticulocytes are less than 80,000/mm3 (80 x 109/L) if the hemoglobin is less than 9 g/dL (90 g/L or 5.6 mmol/L). Monitor for increases in serum creatinine and transaminases. Once the patient has recovered, hydroxyurea may be restarted with a dose that is 2.5 to 5 mg/kg less than the dose associated with the patient s toxicity. Doses then may be increased by 2.5 to 5 mg/kg daily after 12 weeks with no toxicity. 

D. Similar methods were used for modification of the enzymes listed in Table II as well as bovine hemoglobin (see Table III). The choice of conditions for the modification reactions (pH, temperature, etc.) was made mainly based on the properties/stability of each protein. Enzymatic activities were measured by previously reported methods (77,27-25). 

---