Characterization of human residual catalase of an acatalasemic patient by isoelectric focusing and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis followed by electrophoretic blotting and immunodetection

Abstract

Isoelectric points and subunit sizes of catalases in human blood and human cultured skin fibroblasts from acatalasemic and normal subjects were analyzed by isoelectric focusing in agarose gel and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, respectively, followed by electroblotting to polyvinylidene difluoride membranes for immunodetection. The results indicated that the isoelectric point of residual catalase in the C fraction prepared from acatalasemic erythrocytes was identical with that of catalase prepared from normal erythrocytes. The residual catalase in homogenates of acatalasemic cultured skin fibroblasts also reacted with anticatalase rabbit serum and had an isoelectric point identical with that of normal catalase. Subunit sizes of normal and acatalasemic catalases in the C fractions of erythrocytes were also found to be identical on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by electroblotting and immunoenzymatic amplification. The results indicated no substantial difference in molecular size and charge of catalase proteins between normal and acatalasemic erythrocytes and fibroblasts.

Acatalasemia

“…The subunit sizes of normal and acatalasemic catalases in the C fractions prepared from erythrocytes are also identical on SDS-PAGE, followed by electroblotting and immunoenzymatic staining (Ogata et al 1989).…”

“…The isoelectric points and subunit sizes of residual catalases in human blood and cultured skin fibroblasts from Japanese acatalasemic cases and normal Japanese individuals were examined by isoelectric focusing in agarose gel and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by electroblotting onto polyvinylidene difluoride membranes for immunodetection (Ogata et al 1989). The isoelectric point of the residual catalase in the C fraction isolated from hemolysates prepared from acatalasemic erythrocytes on D E A E column chromatography (Thorup et al 1964) is identical to that of catalase prepared from normal erythrocytes (Fig.…”

“…Normal activity (%) Cellular distribution in blood (ratio of activity in reticulocytes to activity in erythrocytes Activity in tissues relative to normal Catalase activity in heterozygotes (hypocatalasemia) Japanese 0.16 _+ 0.02 Slightly higher but not (Ogata et al 1972) significantly different 0.39 _+ 0.23 from normal (1.4-5.6) (Takahara et al 1977) (Ogata et al 1972) Swiss 1.08 + 0.34 Much higher (300-400) (Aebi et al 1964) (Aebi et al 1966) Mutant mice 4.40 +_ 0.30 (Tottori 1987) Moderately higher (25) (Ogata et al 1970) Trace in vermiform appendix (2.7%) Abdominal muscle (UDS) a Cultured skin fibroblasts (2.2%) (Sadamoto 1966) Liver (< 0.01%), placenta (15%), tonsilla (3%) (Aebi et al 1964) Cultured skin fibroblasts (15%) (Krooth et al 1962) Liver (65.7%), kidneys (15.6%), lungs (69.9%) (Tottori 1987) Cultured transformed fibroblast Cell line (30%) (Lewis 1985) Half normal (Takahara et al 1960) Within normal range (Aebi et al 1977) Half normal (Feinstein et al 1966) a UDS, Under detectable sensitivity (Ogata 1989(Ogata ) 1972(Ogata , 1989) (blood, fibroblasts)…”

Characterization of Acatalasemia Detected in Two Hungarian Sisters

“…The residual catalase of the Japanese type does not differ from the normal enzyme with respect to isoelectric point, heat stabil ity and molecular weight [4,5]. On the other hand, in Swiss-type acatalasemia the resid ual catalase activity exhibits a higher mobil-We presented a preliminary report on 2 acatalasémie sisters in Hungary [13].…”