Properties of the catalase molecule obtained from acatalasemic and hypocatalasemic mice Part Ⅲ. Analysis of erythrocyte catalase of acatalasemic mice by DEAE column chromatography and isoelectrofocusing.

Blood samples were taken from the orbital sinus (venous) of normal (C3H/C(sa)C(sa)), acatalasemic (C3H/C(sb)C(sb)) and hypocatalasemic (acatalasemic heterozygote, C3H/C(sa)C(sb)) mice. Packed red cells prepared from mice were washed three times with cold 0.9% NaCl solution to remove the buffy coat. One volume of red cells was hemolyzed with 1.5 volumes of distilled water. The various mouse hemolysates were fractionated into A, B and C fractions by DEAE cellulose column chromatography with a discontinous buffer system. Catalase activity in the eluate was determined by the perborate method, and the eluate was concentrated to 1.0PU/ml. Agarose isoelectric focusing was performed in a pH 3-to-10 Pharmalyte gradient gel at 81℃. The distribution of catalase in the erythrocyte fractions A,B and C from normal and mutant mice were examined.The activity ratio of the C fraction to the total fraction was greater in the descending order of C(sa)C(sa),C(sa)C(sb) and C(sb)C(sb) mice. Catalase of fraction A of C(sb)C(sb), C(sa)C(sb) and C(sa)C(sa) focused to band at pl 6.5-7.3, pl 5.7-6.5 and pl 5.4-5.9, respectively. Similarly, catalase of the fraction C of C(sb)C(sb), C(sa)C(sb) and C(sa)C(sa) focused to a band at pl 6.3-7.2, pl 5.9-6.5 and pl 5.0-6.0, respectively. Thus the isoelectric points of catalases in fractions A and C obtained from the blood of C(sb)C(sb) were higher than those of C(sa)C(sa). The isoelectric points of catalases in fractions A and C of C(sa)C(sb9 were between those of the C(sb)C(sb) and C(sa)C(sa). The results also indicate that the isoelectric point of catalase in fraction A was higher than that of fraction C in each group of mice.