Kinetics of hemoglobin S gelation followed by continuously sensitive low-shear viscosity. Changes in viscosity and volume on aggregation

Steve Kowalczykowski, Jacinto Steinhardt

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

The kinetics of gelation of deoxyhemoglobin S were investigated as a function of temperature, concentration of hemoglobin, and solvent composition. Measurements were made by continuously monitoring the changes in viscosity with time, after polymerization had been induced by rapidly raising the temperature. A specially constructed low-shear viscometer was used. The solution density was also measured continuously to determine whether a volume change accompanied aggregation. The results confirm earlier work in showing that the time-dependence of the viscosity is composed of a variable latent period (several minutes to tens of hours) during which there is only a slight and very gradual increase in viscosity, followed by a stage in which the viscosity rises very sharply within a very short time. The length of the initial latent period is highly dependent upon the HbS ‡ Abbreviations used: Hb, hemoglobin; IHP, inositol hexaphosphate concentration (33rd ± 6 power) and temperature. If the duration is interpreted as the inverse of a reaction rate, the activation energy is 96 ± 10 kcal/mol for solutions containing inosital hexaphosphate. Unlike measurements reported by others, no dependence of the latent period on shear rate was observed at the low shear rate employed. When IHP is omitted from the hemoglobin solutions, qualitatively similar results are obtained; however, the latent period depends on the 26th ± 6 power of the deoxyhemoglobin S concentration and yields an average activation energy of 125 ± 10 kcal/mol. The length of the latent period is increased 40-fold. Tris is known to prevent gelation but the inhibition can be partly reversed by adding IHP. When this is done, highly concentration-dependent latent periods are again observed. The results may be interpreted in terms of nucleation kinetic theories: a critical nucleus composed of approximately 30 hemoglobin molecules is required for gelation; and the energy barrier (which is larger in the absence of IHP) to the formation of this critical aggregate is approximately 100 kcal/mol. Gelation is not accompanied by a detectable volume change (limits 5 × 10-5 g/ml). This indicates that the volume change of the reaction must be less than + 60 cm3/mol when the aggregates represent one half of the HbS available for polymerization.

Original languageEnglish (US)
Pages (from-to)201-213
Number of pages13
JournalJournal of Molecular Biology
Volume115
Issue number2
DOIs
StatePublished - Sep 15 1977
Externally publishedYes

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Sickle Hemoglobin
Viscosity
Hemoglobins
Polymerization
Temperature
Phytic Acid

ASJC Scopus subject areas

  • Virology

Cite this

Kinetics of hemoglobin S gelation followed by continuously sensitive low-shear viscosity. Changes in viscosity and volume on aggregation. / Kowalczykowski, Steve; Steinhardt, Jacinto.

In: Journal of Molecular Biology, Vol. 115, No. 2, 15.09.1977, p. 201-213.

Research output: Contribution to journalArticle

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abstract = "The kinetics of gelation of deoxyhemoglobin S were investigated as a function of temperature, concentration of hemoglobin, and solvent composition. Measurements were made by continuously monitoring the changes in viscosity with time, after polymerization had been induced by rapidly raising the temperature. A specially constructed low-shear viscometer was used. The solution density was also measured continuously to determine whether a volume change accompanied aggregation. The results confirm earlier work in showing that the time-dependence of the viscosity is composed of a variable latent period (several minutes to tens of hours) during which there is only a slight and very gradual increase in viscosity, followed by a stage in which the viscosity rises very sharply within a very short time. The length of the initial latent period is highly dependent upon the HbS‡ ‡ Abbreviations used: Hb, hemoglobin; IHP, inositol hexaphosphate concentration (33rd ± 6 power) and temperature. If the duration is interpreted as the inverse of a reaction rate, the activation energy is 96 ± 10 kcal/mol for solutions containing inosital hexaphosphate. Unlike measurements reported by others, no dependence of the latent period on shear rate was observed at the low shear rate employed. When IHP is omitted from the hemoglobin solutions, qualitatively similar results are obtained; however, the latent period depends on the 26th ± 6 power of the deoxyhemoglobin S concentration and yields an average activation energy of 125 ± 10 kcal/mol. The length of the latent period is increased 40-fold. Tris is known to prevent gelation but the inhibition can be partly reversed by adding IHP. When this is done, highly concentration-dependent latent periods are again observed. The results may be interpreted in terms of nucleation kinetic theories: a critical nucleus composed of approximately 30 hemoglobin molecules is required for gelation; and the energy barrier (which is larger in the absence of IHP) to the formation of this critical aggregate is approximately 100 kcal/mol. Gelation is not accompanied by a detectable volume change (limits 5 × 10-5 g/ml). This indicates that the volume change of the reaction must be less than + 60 cm3/mol when the aggregates represent one half of the HbS available for polymerization.",
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