A critical examination of the reaction of pyridoxal 5-phosphate with human hemoglobin Ao

Biopolymers
M A MariniA Zegna

Abstract

Pyridoxylated normal adult human hemoglobin (HbAo) has been prepared using both oxygenated and deoxygenated HbAo at pH 6.8 and room temperature without the addition of Tris to produce a mixture with P50 of 30 +/- 2 torr and a Hill coefficient of 2.3 +/- 0.1 similar to that of the isolated adult human hemoglobin from the red blood cell. Reduction of the pyridoxylated HbAo in the oxygen-ligated form by sodium borohydride gives unacceptable levels of methemoglobin (i.e., greater than 10%). Excessive foaming and methemoglobin formation can be partially avoided using deoxyHbAo. Reduction with sodium cyanoborohydride is much gentler and gives solutions with less than 5% methemoglobin. Both reducing agents give products with multiple components as shown by analytical chromatography. Radioautography on the isoelectric focusing gels of HbAo treated with 14C pyridoxal 5-phosphate (PLP) shows three major bands for the cyanoborohydride-reduced derivatives and a much more complex mixture of labeled molecules after the sodium borohydride reduction. When pyridoxylated hemoglobin is prepared without reduction, the preparation, after passage through a mixed-bed resin, contains 0.4 equivalents of PLP per heme, and has a P50 of 30 +/- 2 torr and ...Continue Reading

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Citations

Jan 3, 2007·Seminars in Hematology·Robert M Winslow
Aug 24, 2004·Artificial Organs·Robert M Winslow
Jun 17, 2008·Biochimica Et Biophysica Acta·Robert M Winslow

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