Properties of erythrocyte catalase from homozygotes and heterozygotes for Swiss-type acatalasemia

The unstable catalase variant found in the blood of individuals homozygous for Swiss-type acatalasemia and the enzyme species present in heterozygous carriers of this rare defect have been further characterized. The mutant enzyme isolated from acatalasemic red cells is considerably more heat labile and differs in electrophoretic mobility from the normal enzyme. Catalase preparations obtained from heterozygotes consist of an apparently uniform enzyme species, probably representing a molecular hybrid, with properties intermediate to those of the normal and the variant enzyme. However, antigenic identity of catalase from all three sources is observed. Model experiments indicate that hybrid catalase molecules can be produced by recombining normal and variant dimer subunits. Fractionation of erythrocytes according to density and age shows that most of the residual catalase activity is localized in juvenile acatalasemic cells, whereas in normal and heterozygous individuals the catalase activity level does not alter significantly during the life span of the red cells. These findings agree with the observation that there is no gene dosage in heterozygotes, their catalase activity values falling within the normal range.     

Human erythrocyte catalase, isolation with an improved method, characterization and comparison to bovine liver catalase

Catalase enzyme was purified from human erythrocytes. The modified procedure of M?rikofer-Zwez et. al. [Eur. J. Biochem. 11: 49-57, 1969] yielded erythrocyte catalase with high specific activity and with one band on SDS polyacrylamide gel. Its other characteristics (isoelectric point; E405/280, E1%1cm at 280 nm and 405 nm) were in agreement with previously described findings. The results obtained for molecular mass, electrophoretic mobility, chromatographic behaviour on CM-Sepadex gel, addition test, and change of electrophoretic mobility in human serum showed differences between human erythrocyte catalase and bovine liver catalase. These results suggest that human erythrocyte catalase and bovine liver catalase are two distinct catalase forms.     

Spectral studies of human erythrocyte catalase.

The optical absorption and circular dichroic spectra of human erythrocyte catalase (EC 1.11.1.6) and its cyanide, azide, and fluoride derivatives over the wavelength range of 210 to 700 nm are reported. Treatment with acid or alkaline solutions causes spectral changes which may be due to dissociation of the enzyme into subunits and removal of the heme group from the protein. The fractions of the protein structure present as alpha helix, beta pleated sheet, and unordered structure have been estimated from the CD spectrum in the far-ultraviolet region. The CD spectra also indicate that the protein conformation does not change appreciably after cyanide binding. The epr spectroscopy of the native enzyme and its cyanide complex are reported. The spectral results are compared with catalase obtained from other mammalian and bacterial sources.     

The kinetics of cyanide binding by human erythrocyte catalase

The kinetics of the binding reaction of cyanide by human erythrocyte catalase at 25 °C has been studied over the pH range 4.2 to 10.2 by means of temperature jump and stopped flow techniques. Catalase reacts with cyanide at a constant rate in the range pH 4.2 to 8.1 which decreases at higher pH. This is most simply explained by the reaction of catalase with unionized hydrogen cyanide molecules. The pH-independent rate constant for the formation of the catalase-cyanide complex is (1.3 ± 0.1) × 10⁶m^?1 s^?1. The association equilibrium constant and the dissociation rate constant for the catalase-cyanide complex were determined from the relaxation amplitudes of temperature jump experiments and by spectrophotometric titration and are (3.1 ± 0.2) × 10⁵m^?1 and 4.2 ± 0.6 s^?1, respectively in the pH-independent region.     

The partial amino acid sequence of human erythrocyte catalase

More than 95% of the sequence of human erythrocyte catalase (HEC) has been determined. There are at least 41 differences in sequence between it and that of bovine liver and erythrocyte catalases (BLC and BEC). Although the normal subunit length of BLC is 506 residues, BEC has at least 517 and HEC 520 residues. Most differences between HEC and BLC or BEC are conservative substitutions. If the heme-protein contacts as determined by X-ray crystallography are examined, only one of 39 residues in contact with the heme group differs between HEC and BLC or BEC.     

Fractionation and characterization of acidic oligosaccharides and glycopeptides from normal and pathological urines

A general procedure is described for the isolation of urinary acidic oligosaccharides and glycopeptides resulting from catabolism of glycoproteins. This procedure has been applied to normal urine and to urine from patients with diseases of the metabolism, including mucolipidosis and fucosidosis.     

The complete amino acid sequence of bovine liver catalase and the partial sequence of bovine erythrocyte catalase

The data upon which the sequence of the 506 residues in the subunit of bovine liver catalase (BLC) is based are presented in detail. A partial sequence of bovine erythrocyte catalase (BEC) which accounts for 493 residues shows complete concordance with the BLC data. On the other hand, BEC has at least 517 residues, that is, an extension beyond the C terminus of the BLC data. Although normally BLC has only 506 residues, there is evidence that, at some point in its history, it also had the C-terminal extension. It is speculated that this extension is lost in BLC either through a different processing of the molecule in liver than in erythrocytes or by partial degradation in the first stages of catabolism.