Glycolipids from the cell walls of streptococci

A streptococcus, serologically defined as a Z₃III strain was compared with a mutant strain Z₃ lacking the type III polysaccharide antigen. The loss of type antigen represents a decrease in the carbohydrate content of the cell wall of the mutant and is accompanied by long-chain formation, increased sensitivity to streptomycin and agglutination in saline. Cell-wall preparations can be freed of membrane contamination by treatment with hot sodium dodecylsulfate (SDS). This resulted in a doubling of the ratio of muramic acid to lysine and in the disappearance of phospholipids. It could be shown that the membrane-free cell walls of these strains still contained appreciable amounts of glycolipids which could be identified as monoglucosyl glyceride and diglucosyl-diglyceride.     

Reconstitution of the leucine transport system of Lactococcus lactis into liposomes composed of membrane-spanning lipids from Sulfolobus acidocaldarius.

The effect of bipolar tetraether lipids, extracted from the thermophilic archaebacterium Sulfolobus acidocaldarius, on the branched-chain amino acid transport system of the mesophilic bacterium Lactococcus lactis was investigated. Liposomes were prepared from mixtures of monolayer lipids and the bilayer lipid phosphatidylcholine (PC), analyzed on their miscibility, and fused with membrane vesicles from L. lactis. Freeze-fracture electron microscopy demonstrates that the bipolar lipids in the hybrid membranes adopted a monomolecular organization at high S. acidocaldarius lipid content. Leucine transport activity (i.e., delta mu H(+)-driven and counterflow uptake) increased with the content of S. acidocaldarius lipids and was optimal at a one-to-one (w/w) ratio of PC to S. acidocaldarius lipids. Membrane fluidity decreased with increasing S. acidocaldarius lipid content. These data suggest that transport proteins can be functionally reconstituted into membranes composed of membrane-spanning lipids provided that membrane viscosity is restricted.     

Structural and dynamic effects of oxidatively modified phospholipids in unsaturated lipid membranes.

Phospholipid hydroperoxides and phospholipid alcohols are two of the major forms of oxidatively modified phospholipids produced during oxidant stress and lipid peroxidation. The process of lipid peroxidation is known to affect the physiological function of membranes. We, therefore, investigated the effects of lipid peroxidation products on the molecular interactions in membranes. Our study was specifically focused on the effects of lipid peroxidation products on static membrane structure (molecular orientational order) and on the reorientational dynamics of the probe molecules in lipid bilayers. The study was done by performing angle-resolved fluorescence depolarization measurements (AFD) on the fluorescent probe diphenylhexatriene (DPH) and by performing angle-resolved electron spin resonance (A-ESR) measurements on cholestane (CSL) nitroxide spin probes embedded in macroscopically oriented planar bilayers consisting of 2-10% 1-palmitoyl-2-(9/13-hydroperoxylinoleoyl)phosphatidylcholine (PLPC-OOH) or 1-palmitoyl-2-(9/13-hydroxylinoleoyl)phosphatidylcholine (PLPC-OH) in 1-palmitoyl-2-linoleoylphosphatidylcholine (PLPC) or dilinoleoylphosphatidylcholine (DLPC). Both probe molecules have rigid cylindrical geometries and report on the overall molecular order and dynamics. However, being more polar, the nitroxide spin probe CSL is preferentially located near the surface of the membrane, while the less polar fluorescent probe DPH reports preferentially near the central hydrophobic region of the lipid bilayers. The results show that the presence of relatively small amounts of oxidatively modified phospholipids within the PLPC or DLPC membranes causes pronounced structural effects as the molecular orientational order of the probe molecules is strongly decreased. In contrast, the effect on membrane reorientational dynamics is minimal.     

Fermentation of cellulose and production of cellulolytic and xylanolytic enzymes by anaerobic fungi from ruminant and non-ruminant herbivores

Four anaerobic fungi were grown on filter paper cellulose and monitored over a 7–8 days period for substrate utilisation, fermentation products, and secretion of cellulolytic and xylanolytic enzymes. Two of the fungi (N1 and N2) were Neocallimastix species isolated from a ruminant (sheep) and the other two fungi were Piromyces species (E2 and R1) isolated from an Indian Elephant and an Indian Rhinoceros, respectively. The tested anaerobic fungi degraded the filter paper cellulose almost completely and estimated cellulose digestion rates were 0.25, 0.13, 0.21 and 0.18 g · 1^-1 · h^-1 for strains E2, N1, N2, R1, respectively. All strains secreted cellulolytic and xylanolytic enzymes, including endoglucanase, exoglucanase, -glucosidase and xylanase. Strain E2 secreted the highest levels of enzymes in a relatively short time. The product formation on avicel by enzymes secreted by the four fungi was studied. Both in the presence and absence of glucurono-1,5--lactone, a specific inhibitor of -glucosidase, mainly glucose was formed but no cellobiose. Therefore the exoglucanase secreted by the four fungi is probably a glucohydrolase.     

Lipid requirement of the branched-chain amino acid transport system of Streptococcus cremoris

The role of the membrane lipid composition on the transport protein of branched-chain amino acids of the homofermentative lactic acid bacterium Streptococcus cremoris has been investigated. The major membrane lipid species identified in S. cremoris were acidic phospholipids (phosphatidylglycerol and cardiolipin), glycolipids, and glycerophosphoglycolipids. Phosphatidylethanolamine (PE) was completely absent. Protonmotive force-driven and counterflow transport of leucine was assayed in fused membranes of S. cremoris membrane vesicles and liposomes composed of different lipids obtained by the freeze/thaw-sonication technique. High transport activities were observed with natural S. cremoris and Escherichia coli lipids, as well as with mixtures of phosphatidylcholine (PC) with PE or phosphatidylserine. High transport activities were also observed with mixtures of PC with monogalactosyl diglyceride, digalactosyl diglyceride, or a neutral glycolipid fraction isolated from S. cremoris. PC or mixtures of PC with phosphatidylglycerol, phosphatidic acid, or cardiolipin showed low activities. In mixtures of PC and methylated derivatives of PE, both counterflow and protonmotive force-driven transport activities decreased with increasing degree of methylation of PE. The decreased transport activity in membranes containing PC could be restored by refusion with PE-containing liposomes. These results demonstrate that both aminophospholipids and glycolipids can be activators of the leucine transport system from S. cremoris. It is proposed that aminophospholipids in Gram-negative bacteria and glycolipids in Gram-positive bacteria have similar functions with respect to solute transport.     

Splint renal function after captopril in unilateral renal artery stenosis.

The renal extraction ratios of 131I-sodium iodohippurate (131I-Hippuran) and 125I-thalamate were greatly reduced on the affected side by 50 mg captopril in seven out of 14 patients with unilateral renal artery stenosis. With long term captopril 150 mg daily the uptake of 99mTc-diethylenetriaminepenta-acetic acid by the affected kidney, which was determined by scintillation camera renography, became almost zero in these seven patients, indicating severe reduction of the glomerular filtration rate. Function of the affected kidney returned on discontinuing treatment. The reduced extraction of sodium iodohippurate probably reflected a shortened plasma transit time through the kidney due to intrarenal vasodilatation. The reduced extraction of thalamate reflected a low filtration fraction, suggesting that the vasodilatation was, at least in part, at the level of the postglomerular arterioles. Captopril had little effect on the contralateral kidney and on the kidneys of 17 patients with essential hypertension, and serum creatinine concentrations showed minor changes. Radioisotope renography should be performed after beginning captopril treatment in patients with renal artery stenosis. This is also recommended for patients given captopril as a third line drug when renal artery stenosis has not been excluded. Hypertension is these patients is often severe and difficult to control. Renal artery disease is not rare in this difficult group and finding seriously impaired renal function on one side during captopril treatment may be diagnostic.     

The certification of a reference material for the evaluation of the ISO method for the detection of Salmonella

A reference material (RM) containing Salmonella typhimurium was certified as CRM 507 by the Standards, Measurements and Testing Programme of the European Commission. The material consists of a gelatin capsule filled with artificially contaminated milk powder. The material is certified for the evaluation of presence/absence methods based on the ISO 6579 procedure for the detection of Salmonella. In the certification study 11 laboratories determined the presence/absence of Salmonella from each of 50 capsules. They also determined the mean number of colony-forming particles (cfp) and the homogeneity of the batch of RM according to an enumeration procedure. Certified values were calculated for both procedures separately. Based on the presence/absence procedure a fraction of capsules in which no Salmonella could be detected of 2·7% (one-sided 95% confidence upper limit 4·4%) was certified, for the enumeration procedure this fraction was 0·61% (one-sided 95% confidence upper limit 1·6%). The certified mean number of Salmonella cfp in one capsale is 5·9 (two-sided 95% confidence interval 5·3—6·4). Data on the preparation, identification, stability (at storage and higher temperatures) and homogeneity of the material are presented.     

Glycoprotein synthesis in the mucous cells of the vascularly perfused rat stomach. I. Surface mucous cells

We developed a constant-pressure vascular perfusion system of the isolated rat stomach, utilizing an artificial, fluorocarbon (FC-75)-containing medium. Perfusion could be maintained for at least six hours, as demonstrated by the ultrastructure of the mucosal cells and by the constant incorporation of [3H]-galactose in the surface mucous cells. Moreover all mucous cell types in tissue fixed after six hours of perfusion showed the same histochemical reactions for glycoproteins as in tissue fixed shortly after decapitation of the animal. The surface mucous cells of the antrum incorporated 30% less [3H]-galactose, [3H]-serine and [35S]-sulphate than those of the fundus. The amount of radioactivity incorporated per cell did not decrease during a subsequent 2 hour chase.