Carbohydrate Composition of Variant-Specific Surface Antigen Glycoproteins from Trypanosoma brucei

SYNOPSIS. The carbohydrate of variant-specific surface antigen glycoproteins from bloodstream forms of 13 cloned variants of Trypanosoma brucei was analyzed by gas-liquid chromatography. The glycoproteins contained from 6 to 17% carbohydrate by weight, and all contained the same 4 sugars: mannose, galactose, glucose, and glucosamine (probably as N-acetyl-glucosamine). The glycoprotein from variant 048, strain 427 contained (±20%) 11 mannose, 4 galactose, 4 glucose, and 5 glucosamine residues/mole of glycoprotein (molecular weight 65,000). Glucose was an integral component of the glycoproteins, not dissociable by sodium dodecyl sulphate, 8 M urea, or 1 M acetic acid. Some of the glucose was dissociated by trichloroacetic acid. Most of the glycoproteins formed precipitin bands with concanavalin A in Ouchterlony double diffusion, but none formed such bands with wheat germ agglutinin or Ricinus communis lectin (molecular weight 120,000).     

Transference of the high molecular weight carbohydrate sequences of fetuin to the surface of carrot embryo protoplasts

A method is described for the removal of the carbohydrate sequences of glycoproteins, and their covalent attachment to hydrocarbon chains. These synthetic membrane components may then be incorporated into liposome and cell membranes. Pronase-liberated glycopeptides derived from fetuin were linked by a reduced Schiff's base linkage to tetradecyl aldehyde. The resulting glycolipid was incorporated by external addition, into phosphatidylcholine liposomes. Glycolipid transfer to these liposomes rendered them suseptible to agglutination by wheat germ lectin, which binds N-acetylneuraminic acid, the terminal carbohydrate of the high molecular weight fetuin sugar sequence. Sequential removal of the terminal sugars, and subsequent agglutination behaviour towards various lectins, suggests that the carbohydrate sequence had been transfered intact. The glycolipid was incorporated into plant protoplast membranes by incubation with glycolipid-containing liposomes for 2 h at 37°C. These synthetic glycolipids may find a use in the study of carbohydrate-based recognition systems in animal and plant membranes. In addition they may prove useful in the development of cell and membrane tagging and handling techniques, by the insertion of sugar groups not normally present in these membranes.     

Adsorption of non-membrane proteins on the surface of model phospholipid membranes

1. Two new methods are proposed for enhancement of the binding of hydrophilic proteins by liposomes. 2. An alkylating derivative of phosphatidic acid has been obtained by its reaction with N,N,N′-tris(2-chloroethyl)-N′- (p-formylphenyl)propylene-1,3-diamine. The alkylating activity of this derivative is very low due to the electron-acceptor effect of the formyl residue. Phosphatidylcholine liposomes which contain this alkylating derivative in the lipid bilayer may be obtained. The compound residing in the outer monolayer may be reduced by NaBH₄. Upon reduction, the formyl residue is transformed into a hydroxymethyl residue. Therefore, the alkylating group of the compound is activated, and proteins may be attached covalently to the outer monolayer by alkylation with such chemically reactive liposomes. 3. Reaction of alkylating liposomes with myoglobin results in covalent binding of this hydrophilic protein. Complement-mediated leakage of such myoglobin-carrying liposomes may be induced by antibodies against myoglobin. 4. Modification of hydrophilic proteins with dansyl chloride results, even at small extents of modification, in a dramatic increase of the affinity of such proteins to phosphatidylcholine liposomes.     

A technique for the long-term application of surface electrodes

A technique is reported for the long-term application of surface electrodes for ambulatory electromyographic (EMG) recording. Prior to electrode application the surrounding skin is lightly painted with tincture of benzoin. This treatment improves adherence to the skin of disposable electrodes and electrode attachment collars, reduces skin trauma associated with electrode removal, and minimizes sensitivity to electrode adhesives.This research was supported in part by NIH grant No. NS25114.     

Surface heterogeneity of rat sperm during maturation

Rat sperm isolated from the caput and caudal epididymis and the vas deferens were subjected to multiple partition in aqueous two-phase systems. The technique was used to reveal heterogeneity of a sperm population with respect to particular surface properties. Sperm from all three regions gave broad distributions indicative of heterogeneous cell populations. Greatest heterogeneity was observed for cauda sperm with caput and was sperm producing similar distributions.Following multiple partition sperm from different regions of the distribution profiles were immunostained with three antibodies known to recognise maturation antigens. The results show that some antigens are acquired during epididymal transit whilst others are present throughout.The partition (surface heterogeneity) seen cannot therefore be explained solely by the distribution of the antigens recognised by 2D6, 6B2 and 3D5.     

Structural features of archaebacterial cell envelopes

Regularly arrayed surface (glyco)proteins—often referred to as S layers—are a common feature of the cell envelopes of almost all archaebacteria. We have selected some examples (Halobacterium, Sulfolobus, Thermoproteus, Pyrobaculum, Staphylothermus), and we describe the structure of their surface layers as revealed primarily by electron crystallography. In spite of a considerable diversity in shapes and dimensions, some common structural features emerge from the comparison. The glycoprotein arrays are composed of oligomeric units which are anchored in the plasma membrane; extended spacer or linker domains maintain the bulk of the more or less porous surface layers at a constant distance above the membrane surface, thus creating a quasi-periplasmic compartment. Functions ascribed to surface layers, such as compartmentalization, shape maintenance and determination, and adhesion are discussed.     

SEM observations on polar body elimination and changes of the oocyte surface during egg maturation in the newt Triturus alpestris (Amphibia,Urodela)

Summary We have studied the surface of the animal half of ovulated newt eggs recovered from different portions of the oviduct. The germinative area, about 40 m diameter, is localized in the region of the whitish polar spot, about 450 m diameter. The structural changes in the germinative area are connected with the formation and extrusion of the first and second polar bodies. Of the two types of oviductal eggs observed, those covered with microvilli (type 1) were found only in the ostial portions of the oviduct, whilst those covered with microfolds (type 11) were found more distally. The structural difference between these two types may be related to the known reduction in surface area of the cell membrane during oocyte maturation. Offprint requests to: W. Kilarski     

Application of the functional-form model to the culture of seaweeds

Selecting the most appropriate species or strains is an important first step in the development of most algal cultivation systems and is usually a tedious, time-consuming, and expensive step. The functional-form model, first developed to synthesize the adaptive significance of easily assessed thallus-form attributes relative to the productivity and survival of benthic macroalgae, is applicable to the culture of seaweeds and can expedite species or strain selection. The production ecology aspects of the model are useful particularly for applications where the desired product is not species-specific, e.g., systems in which the emphasis is on algal production, such as algal biomass farms and wastewater treatment. A thallus-form with a high surface area: volume ratio is more suited for rapid production and nutrient uptake. The utility of this model to strain selection is demonstrated with the red alga Gracilaria tikvahiae, a species that has been considered a maricultural candidate for a number of utilizations. A continuum of surface area: volume ratios for eight clones of G. tikvahiae showed that this ratio decreased as morphological complexity increased and was a good predictor of both short-term photosynthesis and long-term growth rate. Clones near opposite ends of the surface area: volume ratio spectrum had significant differences for both photosynthesis and growth. Each clone of G. tikvahiae possesses concomitant combinations of benefits as well as costs, which should be carefully evaluated for the cultivation application of interest. Knowledge of functional-form relationships in seaweeds can significantly expedite their successful cultivation.     

The surface coat of bloodstream forms of Trypanosoma musculi from mice

Iodination and immunoprecipitation techniques together with indirect fluorescent antibody tests identified two polypeptides (SP) of molecular weights 88,000–92,000 and 66,000–70,000 in the surface coat of bloodstream forms of the mouse trypanosome, Trypanosoma musculi. As parasites multiply and enter the early plateau phase of infection the 88,000–92,000 SP is present while the 66,000–70,000 SP is only detectable after the mid-plateau phase. Western blotting of parasite extracts showed that the 88,000–92,000 SP was present throughout the course of infection, but it appears to become masked by the 66,000–70,000 SP or possibly immunoglobulin from about 16 days after infection. Based on results when Western blots of parasite extracts were probed with antibodies affinity purified against the 88,000–92,000 SP, the two SP appear to be immunologically related and the smaller may be a cleavage product of the larger. This would explain why affinity purified antibodies to each SP bound to trypanosomes collected 8 days after infection, when only the 88,000–92,000 is detectable in parasite extracts. However, the failure of antibodies affinity purified against the 66,000–70,000 SP to bind to the 88,000–92,000 SP in Western blots suggests that the smaller SP has some epitopes that are immunologically distinct from those of the larger SP.     

Stage-specific cuticular proteins of Ostertagia circumcincta and Ostertagia ostertagi

In this study we have shown that NHS-biotin and I¹²⁵-streptavidin can detect cuticular polypeptides of Ostertagia spp. The labelled polypeptide profile of intact nematodes is simple compared to the profile obtained by labelling homogenates. None of the major internal polypeptides are labelled and the subset of proteins labelled in intact nematodes appears to be mainly surface associated. The results presented here demonstrate that NHS-biotin may be used as a reagent for the analysis of surface polypeptides. The surface polypeptide profiles of the five major developmental stages (L1, L2, L3, L4 and adult) of Ostertagia circumcincta show a series of stage-specific molecules with no polypeptides common to all stages, indicating that the cuticle is a dynamic structure which changes throughout the life cycle. Similarity comparison of Ostertagia ostertagi L3 and L4 stage surface profiles showed that each stage is clearly distinct; comparison of these stages between the two species shows an overall similarity.