Separation of asymmetrical hybrid containing hemoglobin F by anaerobic anion-exchange high-performance liquid chromatography

✍ Scribed by Charlotte Yu Ip; Toshio Asakura

• Publisher: Elsevier Science
• Year: 1986
• Tongue: English
• Weight: 449 KB
• Volume: 156
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(86)90264-2

✦ Synopsis

Asymmetrical

hybrid hemoglobins formed from mixtures of oxyhemoglobins S and F and A and F were separated by high-performance liquid chromatography on a 4.6 X 250 mm wide-pore polyethyleneimine-silica gel column under anaerobic conditions. The resulting HPLC chromatogram showed three peaks, with the middle peak representing the hybrid hemoglobin.

The areas ofthese three peaks were quantified and the amount of hybrids formed was less than that predicted theoretically.

We found that the deviation was due to the equilibrium constant of the FS hybrid hemoglobin differing from that ofthe parent hemoglobins.

In this report. we introduce the anaerobic recycle ion-exchange HPLC method to determine the rate of dissociation of AS and FS hybrid hemoglobins at constant pH buffer conditions. The results obtained by this method demonstrate that FS hybrid hemoglobin is more unstable than AS hybrid hemoglobin.

The free energy of association for asymmetrical hybrids containing hemoglobin F is approximately 0.6 Kcal/mol greater than that of the symmetrical parent hemoglobins.