Characteristics and amino-acid composition of a c-type cytochrome in electron acceptor function during thiosulfate-linked photoautotrophic growth of Rhodopseudomonas palustris.

The purple bacterium Rhodopseudomonas palustris (Rhodospirillaceae) was grown in the light with thiosulfatee as the only electron source and HCO theta 3/CO2 as carbon requirement. During thiosulfat oxidation, photolithoautotrophically growing cells transferred the electrons enzymatically towards an endogenous, soluble cytochrome of type c. The cytochrome c in electron acceptor function was purified to homogeneity and appeared as a single protein band in a dodecyl sulfate disc gel electrophoresis. Its molecular mass was determined to be about 16000 Da and its pI value 10.0. The determination of its amino-acid composition revealed a long-chained cytochrome represented by more than 120 amino-acid residues with a characteristic content of lysine and a lack of tryptophan.

Age-related changes in the composition of proteins in the trabecular meshwork of the human eye.

The composition of the trabecular meshwork proteins of human eyes ranging in age from 36 days to 84 years was examined by polyacrylamide gel electrophoresis and amino acid analysis. Proteins of different molecular weights could be extracted from the tissue with acetic acid. Although their electrophoretic patterns became less distinct with increasing age, proteins of molecular weights ranging from 50 000 to 69 000 always prevailed. The amino acid compositions of the acetic acid-insoluble trabecular meshwork residues revealed the prevalence of collagenous proteins. The peptide maps produced by treatment with cyanogen bromide indicate that most of the fragments solubilized from the trabecular meshwork of younger eyes are derived from type I collagen. Beyond 40 years of age, the trabecular meshwork was resistant to cyanogen bromide and pepsin digestion. A rough estimate of the distribution of collagen types in the trabecular meshwork was based on 3-hydroxyproline/4-hydroxyproline ratios, indicating an age-related increase of type I collagen from about 55 to 70 per cent, and of type IV collagen from about 2 to 5 per cent of the total protein present. During ageing, some of the protein-bound methionine is oxidized to methionine sulfoxide, reaching about 35 per cent of the total methionine content at the age of 20 years and, with a slower rate of oxidation, a mean value of 40 per cent at 80 years of age. Electron-microscopic analysis of specimens remaining undissolved after cyanogen bromide cleavage and pepsin treatment no longer revealed regular collagenous fibrils but rather elastic-like fibers surrounded by wide sheaths consisting of fine fibrils with a regular cross-banding periodicity of 40-50 nm. In addition, clusters of so-called curly (lattice) collagen were found. The amino acid composition of this insoluble material suggests that altered collagen-like molecules prevail among the proteins of the residues.

Molecular characterization of glutamine synthetase from the nitrogen-fixing phototrophic bacterium Rhodopseudomonas palustris.

The phototrophic bacterium Rhodopseudomonas palustris assimilated ammonium via glutamine synthetase and glutamate synthase. Diazotrophic and ammonium-grown cells had high levels of both enzymes, whereas enzymes of alternative assimilatory pathways were absent or had only low activities. Glutamine synthetase was purified to electrophoretic homogeneity within three steps by dye-ligand and ion exchange chromatography. Electron microscopy revealed a dodecameric molecular entity which was in accordance with parameters derived from electrophoretic techniques. The molecular weight of the enzyme monomer was 558000; that of the dodecamer 670000. The amino acid composition of R. palustris glutamine synthetase was determined and compared by a statistical method with other known enzyme compositions from prokaryotic and eukaryotic origins.

[Formation of high-molecular aggregates between serum amylase and colloidal plasma substitutes (author's transl)]. / Die Bildung hochmolekularer Komplexe aus Serumamylase und kolloidalen Plasmaersatzmitteln.

The distribution of molecular weights of human serum amylase was studied by gel filtration of serum, obtained before and after infusion of the colloidal plasma substitutes hydroxyethyl starch, dextran and gelatin, respectively. Both in serum, drawn after intravenous infusion of hydroxyethyl starch, and in a solution of hydroxyethyl starch with serum we observed a significant increase in the molecular weight of serum amylase. The occurrence of this "macroamylase" may be explained by the formation of aggregates between hydroxyethyl starch and amylase. Because of its high molecular weight the elimination of this, presumably, enzyme-substrate-complex is retarded, thus leading to the observed increase of serum amylase activity. In contrast to these observations concerning hydroxyethyl starch no change in the apparent molecular weight of serum amylase was observed following the infusion of either gelatin or dextran or their solutions with serum.

Hydroxyethyl starch-induced macroamylasemia.

Summarizing, the following results were obtained: 1. After infusion of 500 ml hydroxyethyl starch 6% in all patients an increase of serum amylase occurred, which in most cases reached values twice as high as the basal value. 2. This hyperamylasemia is caused by the formation of a high molecular HES-amylase complex, which cannot be easily eliminated. 3. In no case was hyperamylasemia associated with other clinical signs or symptoms.

Macroamylasaemia after treatment with hydroxyethyl starch.

After infusion of 500 ml of 6% hydroxyethyl starch into fifty-four patients an increase of serum amylase was observed which in fifty-one cases exceeded the upper limit of normal (190 U/l). In most cases serum amylase reached twice the basal value. Renal function influenced the duration of the increase in serum amylase, but not the maximum increase (201+/-15 U/l; mean+/-SEM). In patients with advanced renal failure (glomerular filtration rate (GFR) = 2-10 ml/min) serum amylase was still markedly elevated after 72 h (298+/-24 U/l; mean+/-SEM). In patients with normal renal function (GFR greater than 90 ml/min) serum amylase decreased to 183+/-40 U/l (mean+/-SEM) within 72 h without reaching basal values. After infusion of HES no changes were observed in serum lipase or in amylase or lipase activities in duodenal secretion. Amylase excretion in the urine decreased. The assumption of a macroamylasaemia caused by formation of an HES-amylase complex was confirmed by gel filtration. The elimination from plasma of this high molecular enzyme-substrate complex is slow and causes hyperamylasaemia. In no case was the macroamylasaemia associated with signs or symptoms. An awareness of this causal relationship seems to be important, to avoid the erroneous diagnosis of a pancreatic disease.

Congenital dyserythropoietic anaemia, types I and II: aberrant pattern of erythrocyte membrane proteins in CDA II, as revealed by two-dimensional polyacrylamide gel electrophoresis.

The protein composition of the erythrocyte membrane of two patients with CDA I and four patients with CDA II has been investigated by two-dimensional polyacrylamide gel electrophoresis. The two-dimensional patterns of erythrocyte membrane proteins of the patients with CDA were compared with the well-established pattern of normal individuals. No alterations could be detected in the patients with CDA I, but a striking deviation from normal was observed in all patients with CDA II. The major aberrations of the erythrocyte membrane proteins in CDA II are the lack of protein B1.1, the drastically increased concentration of protein B1.4 and the presence of a new component, C2'. These results are discussed in relation to the known morphological and immunological abnormalities of the erythrocyte membrane in CDA II.

Two-dimensional polyacrylamide-gel electrophoresis of the proteins and glycoproteins of the human erythrocyte membrane.

A two-dimensional polyacrylamide gel electrophoresis technique has been developed, improving the analytical separation of some proteins and glycoproteins of the human erythrocyte membrane. Freshly prepared membranes are totally solubilized, subjected to dodecylsulfate--polyacrylamide gel electrophoresis in the first dimension, followed by electrophoresis in the second dimension, using a detergent-free polyacrylamide gradient gel. By this method the proteins of the human erythrocyte membrane could be resolved into a two-dimensional pattern, which has been shown to be highly reproducible with respect to various blood-groups and within one blood-group from specimen to specimen. The method enables especially the investigation of the hydrophobic and very likely integrated membrane proteins and glycoproteins. Thus, band III[Fairbanks, G., Steck, Th. & Wallach, D. F. H., Biochemistry, 10, 2606--2617 (1971)] could be shown to consist of five proteins, one of them being the major glycoprotein of the human erythrocyte membrand. The two spectrin bands differed considerably in their two-dimensional patterns. The value of the given method for the investigation of membrane defects, which may be linked with various diseases of human erythrocytes, could be demonstrated in the case of two patients suffering from congenital dyserythropoetic anaemia.

Zur primärstruktur des cytochromes c des steppenzebras (Equus quagga Boehmi).

Cytochrome c of the zebra (Equus guagga Boehmi) differs from horse cytochrome c in having in its polypeptide chain one serine residue instead of none and 9 threonine residues instead of 10. This replacement has been localised at position 47 of the sequence.