Comparative immunological and electrophoretic analysis of Gc protein in sera from various animals

1. On immunodiffusion, using an anti-human Gc antibody, serum Gc in all mammals tested revealed a partial identity with human Gc. 2. The relative cross-reactivities of serum Gc in monkeys, dogs, cats and rats with human Gc antiserum were found to be more than 70% while the serum Gc in other mammals (pigs, cattle, goats and a guinea pig) was less than 50%. 3. Testing, using the isoelectrofocusing method, showed specific patterns of Gc in the mammals. In the sera of cats and cattle, Gc polymorphisms were detected. 4. Neuraminidase treatment affected the isoelectrofocusing Gc patterns of pigs, goats and cattle, whereas those in other mammals remained unchanged.     

Cross-linking of elongation factor 2 to rat-liver ribosomal proteins by 2-iminothiolane

Complexes containing rat liver 80S ribosomes treated with puromycin and high concentrations of KCl, elongation factor 2 (EF-2) from pig liver, and guanosine 5'-[beta, gamma-methylene]triphosphate were prepared. Neighboring proteins in the complexes were cross-linked with the bifunctional reagent 2-iminothiolane. Proteins were extracted and then separated into 22 fractions by chromatography on carboxymethylcellulose of which seven fractions were used for further analyses. Each protein fraction was subjected to diagonal polyacrylamide/sodium dodecyl sulfate gel electrophoresis. Nine cross-linked protein pairs between EF-2 and ribosomal proteins were shifted from the line formed with monomeric proteins. The spots of ribosomal proteins cross-linked to EF-2 were cut out from the gel plate and labelled with 125I. The labelled protein was extracted from the gel and identified by three kinds of two-dimensional gel electrophoresis, followed by autoradiography. The following proteins of both large and small subunits were identified: L9, L12, L23, LA33 (acidic protein of Mr 33000), P2, S6 and S23/S24, and L3 and L4 in lower yields. The results are discussed in relation to the topographies of ribosomal proteins in large and small subunits. Furthermore we found new neighboring protein pairs in large subunits, LA33-L11 and LA33-L12.     

Increase in negative supercoiling of plasmid DNA in Escherichia coli exposed to cold shock

Negative supercoiling of plasmid DNA in Escherichia coli cells can decrease transiently when exposed to heat shock. The effect of cold shock on DNA supercoiling was examined, and analysis by agarose gel electrophoresis in the presence of chloroquine revealed that negative supercoiling of plasmid DNA in cells increased when cells were exposed to cold shock. This increase was transient and was nil when the cells were pretreated with nalidixic acid, an inhibitor of DNA gyrase. In a mutant deficient in expression of HU protein, the increase in negative supercoiling of DNA by cold shock is less apparent than in wild-type cells. It is proposed that DNA gyrase and HU protein have a role in the DNA supercoiling reaction seen with cold shock.     

Involvement of endogenous digitalis-like substance in genesis of deoxycorticosterone-salt hypertension

In an attempt to evaluate the role of endogenous digitalis-like substance in the genesis of deoxycorticosterone (DOCA) -salt hypertension, the effects of intravenous anti-digoxin antibody on the blood pressure response were observed in male Wistar rats that underwent heminephrectomy followed by treatment with DOCA and saline. Administration of anti-digoxin antibody caused a marked decrease in the blood pressure which continued for about an hour. Such a change in the blood pressure was not observed in pertinent control animals. Thus, it seems that endogenous digitalis-like substance plays an important role in the genesis of DOCA-salt hypertension.     

Morphological Changes during Conversion of Clostridium saccharoperbutylacetonicum to Protoplasts by Sucrose-Induced Autolysis

When exponentially growing cells of Clostridium saccharoperbutylacetonicum (ATCC 13564) were exposed to hypertonic concentrations of sucrose (0.3–0.5 M), rapid degradation of the cell wall occurred (sucrose-induced autolysis). The morphological changes from the original rod-shaped cells to protoplasts during the sucrose-induced autolysis were investigated by phase contrast and electron microscopy. When the cells were autolysed in the sucrose solution (0.35 M), each cell began to swell at the middle or at one pole and then formed a small bulb at the swollen part. The bulb consisted of the cytoplasm which was enveloped by the plasma membrane and extruded from the small gap produced by the degradation of the cell wall. The bulb gradually enlarged as lysis progressed, and finally became a protoplast which had no cell wall. The large pre-division cell frequently formed the bulb at the middle (septal site), while the small post-division cell formed the bulb at the pole.     

Fatty Acid Hydroxamate Formation and Ester Hydrolysis by Cell-free Extracts of Micrococcus sp.

When Micrococcus sp. which was isolated from soil assimilated azelaic acid as a sole carbon source, cell-free extract of the organism catalyzed enzymic fatty acid hydroxamate formation. The enzyme was effective only for mono-carboxylic acid, but not for di-carboxylic acids such as azelaic acid. The activity was high with higher fatty acid such as oleic acid. Some of the properties of higher fatty acid hydroxamate formation were investigated.

Olelylhydroxamate was formed with the high concentration of hydroxylamine. The reaction was inhibited by PCMB, but recovered by the addition of SH-compounds (such as cysteine).

On the other hand, when methylacetate was used as a sole carbon source and cell-free extract of Micrococcus sp. hydrolyzed several fatty acid esters. The fatty acid hydroxamate degradation by esterolysis are also discussed.     

Properties of residual catalase in the erythrocytes of Japanese-type acatalasemia

Summary In Japanese-type acatalasemia erythrocytes, the presence and properties of residual catalase were determined and compared with those of normal erythrocyte catalase. Residual catalase activity was proved by titration, active staining after polyacrylamide gel electrophoresis, and measurement of oxygen evolution. Residual catalase protein, demonstrated by double immunodiffusion, was similar to that of normal catalase. The properties of residual catalase activity were identical with those of normal catalase activity. It occurred as three fractions of equal specific activity by DEAE column chromatography. These observations suggest that Japanese-type acatalasemia contains residual catalase with properties similar to those of normal catalase.