Transbilayer distributions of red cell membrane phospholipids in unilamellar vesicles

The phospholipid organization in unilamellar vesicles comprised of various purified phospholipid components of monkey erythrocyte membrane was ascertained using phospholipase A2 and trinitrobenzenesulfonic acid as external membrane probes. The vesicles were formed by sonication or detergent dialysis and fractionated by centrifugation or gel permeation chromatography. Experiments were done to confirm that the phospholipase A2 treatments did not cause lysis or induce fusion of the vesicles. This enzyme hydrolysed only the glycerophospholipids in the outer surface of the vesicles. The amounts of the external phospholipids determined by this enzymatic method were verified using the chemical probe, trinitrobenzenesulfonic acid. The choline-containing phospholipids and phosphatidylethanolamine localized randomly in the two surfaces of sonicated vesicles (outer diameter, about 30 nm), whereas phosphatidylserine preferentially distributed in the inner monolayer. This phosphatidylserine asymmetry virtually disappeared in detergent dialysed vesicles (outer diameter, about 45 nm). Furthermore, inclusion of cholesterol in both the types of vesicles resulted in more random glycerophospholipid distributions across the plane of vesicles bilayer, presumably due to the cholesterol-induced increases in the size of vesicles. These results demonstrate that the transbilayer distribution of erythrocyte membrane phospholipids in unilamellar vesicles are controlled mainly by the surface curvature rather than by interlipid interactions, and therefore suggest that phospholipid-phospholipid and phospholipid-cholesterol interactions should not play any significant role in determining the membrane phospholipid asymmetry in red cells. It is proposed that this asymmetry primarily originates from differential bindings of phospholipids with membrane proteins in the two leaflets of the membrane bilayer.     

Metalloproteinase inhibitors from bovine cartilage and body fluids

Inhibitors of the mammalian metalloproteinases, collagenase, proteoglycanase and gelatinase were isolated from bovine cartilage (extracts and culture medium) and bovine amniotic fluid and serum. These inhibitors either bind or do not bind to concanavalin-A--Sepharose, with Mr (gel filtration) of about 30 000 and 20 000, respectively. Cartilage and chondrocyte culture media contained only concanavalin-A-binding inhibitors whereas cartilage extracts contained only a non-binding inhibitor: serum and amniotic fluid contained both forms of inhibitory activities. In moist biochemical respects, particularly in their abilities to inhibit metalloproteinases, all of the inhibitors were found to be similar. It is concluded that the forms of the inhibitors that differ in Mr may be closely related to the tissue inhibitor of metalloproteinases (TIMP) previously purified from rabbit and human sources. These findings help to clarify other studies on collagenase inhibitors and support the concept that TIMP-like inhibitors may be important in the control of connective tissue degradation.     

Target antigens in malaria transmission blocking immunity

Malaria transmission blocking immunity has been found to operate against two distinct phases of development of malaria parasites in the mosquito midgut: (i) against the extracellular gametes and newly fertilized zygotes shortly after ingestion by a mosquito of parasitized blood and (ii) against the zygotes during their subsequent development into ookinetes. Immunity is antibody-mediated and stage-specific. A set of three proteins, synthesized in the gametocytes, expressed on the surface of the gametes and newly fertilized zygotes and subsequently shed during later transformation of the zygotes, has been identified as the target antigens of anti-gamete fertilization blocking antibodies. A single protein, synthesized and expressed on the zygote surface during its development to ookinetes, has been identified as the target of antibodies which block the development of the fertilized parasites in the mosquito. Immunization of human populations against gamete or zygote antigens, while not directly protecting an immunized individual from inflection, would reduce the transfer of malaria within the population. Such immunity, in addition to reducing the overall rate of malaria transmission, would, if combined with a vaccine against the asexual (disease-causing) stages, reduce the chance of selection of parasites that are resistant to the asexual vaccine by preventing their entry into the mosquito population.     

The apical control of lateral bud development in excised shoot tips of Cuscuta reflexa cultured in vitro

Excised shoot tips of Cuscuta reflexa Roxb. (dodder), a rootless and leafless angiospermic plant parasite, were cultured in vitro for the study of the control of lateral bud development by the apex. In a chemically defined medium lacking hormones, the basal bud alone developed into a shoot. The addition of coconut milk to the growth medium induced the activation of multiple lateral buds, but only a single bud developed further into a shoot. The decapitation of this shoot induced the development of another shoot and the process could be repeated. This showed the controlling effect of the apex in correlative control of bud development. Application of indole-3-acetic acid to the shoot tip explant delayed the development of the lateral bud. Gibberellic acid A₃ induced a marked elongation growth of the explant and reinforced apical dominance. The direct application of cytokinin to an inhibited bud relieved it from apical dominance. A basipetally decreasing concentration gradient of auxin may prevail at the nodes. Bud outgrowth is probably stimulated by cytokinin produced locally in the bud.     

Axial ranking of residues in collagen in relation to edge association and fibril formation

In our earlier analysis of intermolecular interactions between collagen molecules, a major concern with the program employed is that it compared numbers of interactions between residues located on edges of defined, identical width and thus would not necessarily compare the same number of residues in each edge. This would be particularly true of some values of θ where well-defined vertical ranking of residues occurs. We have examined ranking of residues in relation to intermolecular edge association between bovine skin [α1(I)]₃ model collagen molecules by utilizing two different methods of counting intermolecular interactions between residues. The interaction peaks at θ = 27.69° and 36.00° are absent or relatively less intense in the plots obtained by utilizing radial distances between interacting residues instead of vertical bands of defined width. These studies suggest caution in accepting recently reported analyses of superhelix coiling of the collagen molecule which point to values of 27.69° or 36.00° for the twist of the superhelix. Although intramolecular interactions clearly point to interaction of collagen molecules at D intervals, they are insufficiently restricted in distribution to provide a reliable estimate of the superhelix angle by procedures so far employed.     

Proleptonchoides southindiae n. gen., n. sp., a New Leptonchoid from South India

Proleptonchoides southindiae n. gen., n. sp. (Dorylaimida: Leptonchidae), is described from soil around false tobacco (Lobelia excelsa) and cardamom (Elettaria cardamomurn) in South India. P. southindiae is prodelphic, has a short constricted esophageal bulb and flanged odontophore, and is phylogenetically close to Proleptonchus.     

Effect of boron application on barley in a sierozem sandy soil

Summary In a pot culture experiment on a sierozem sandy soil (pH 8.2) rates of added B at 3 ppm although decreased root yield significantly but shoot and grain yield was unaffected even at 6 ppm added B, even though shoot B concentration was as high as 360 ppm and Ca/B ratio as low as 11. At 6 ppm applied B, shoot yield was increased by 18.5 per cent, whereas grain yield was at par with control. The results suggested that Ca/B ratio in barley straw was not a reliable index for determing the magnitude of B problem in the soil.     

Structure and function in galactosyltransferase. Sequence locations of alpha-lactalbumin binding site, thiol groups, and disulfide bond

The region(s) of bovine galactosyltransferase that interacts with the lactose synthase regulatory protein alpha-lactalbumin was investigated using trace 3H acetylation to probe the effects of alpha-lactalbumin on the reactivities of the individual amino groups of galactosyltransferase. In the presence of Mn2+, alpha-lactalbumin was found to reduce the reactivities of lysines 93 and 181 and to increase the reactivities of one or more of lysines 230, 237, and 241. The addition of N-acetylglucosamine (20 mM), which enhances complex formation between the two proteins, did not significantly alter the pattern of perturbation. These results indicate that the NH2-terminal region of the catalytic domain of galactosyltransferase, and possibly part of the proline-rich "stem" region, is affected by the association with alpha-lactalbumin and is therefore implicated in the binding of acceptor substrates. In a separate study only cysteines 176, 266, and 342 of galactosyltransferase were found to react with [3H]iodoacetic acid under denaturing conditions. From their lack of reactivity it is deduced that the remaining two cysteines, residues 134 and 247, are joined in a disulfide linkage. From these results and those of a previous study of UDP-galactose binding (Yadav, S., and Brew, K. (1990) J. Biol. Chem. 265, 14163-14169) it appears that the soluble form of galactosyltransferase is composed of two domains, the NH2-terminal 150 residues containing the Cys134-Cys247 disulfide bond, which functions in alpha-lactalbumin and acceptor binding, and the COOH-terminal region, which is involved in UDP-galactose binding.