A reassessment of the use of herbicide binding to measure photosystem II reaction centres in plant thylakoids

The binding of the herbicide atrazine to thylakoid membranes is often used to quantify Photosystem II reaction centres. Two atrazine binding sites, with high and low affinities, have been observed on the D1 and D2 polypeptides of Photosystem II, respectively (McCarthy S., Jursinic P. and Stemler A. (1988) Plant Physiol. 86S:46). We have observed that the accessibility of the low-affinity binding sites is variable, being limited in freshly isolated thylakoids or in fresh frozen-thawed thylakoids, but increasing during storage of the membranes on ice. In contrast, the accessibility of the high-affinity binding sites, which are titratable at low concentrations (< 500 nM) of herbicide, is much less variable, although the dissociation constant is greatly influenced by ethanol. We conclude that to quantify Photosystem II reaction centres by atrazine binding, it is sufficient and more reliable to assay only the high-affinity binding sites.     

Genetic variation in natural and laboratory populations of the marsupial Sminthopsis crassicaudata

Gel electrophoresis of blood proteins has detected allelic variation at five loci (TRF, PGD, SOD, ADA, GPI) in a laboratory colony of the dasyurid marsupial Sminthopsis crassicaudata. Family data show no significant departures from Mendelian expectations. Analysis of blood from wild-caught progenitors of the colony revealed significant differences in gene frequency between groups of animals captured from different parts of southern and central Australia and showed that there are two major population clusters. These interpopulation differences are particularly marked at the TRF locus and indicate that the river Murray is a barrier for this species.This project was supported by a grant from the Australian Research Grants Scheme.     

β-d-Glucosidases and related enzymic activities in pig kidney

1. The beta-glucosidase activity of pig kidney is located in the unsedimentable fraction of the cell and is not associated with the lysosomes. 2. The enzyme is active towards beta-d-glucosides, beta-d-galactosides, beta-d-xylosides and alpha-l-arabinosides. 3. These activities could not be separated by gel electrophoresis, gel filtration or DEAE-cellulose chromatography. 4. Response to inhibitors, heat-denaturation and competitive substrates suggests that a single active site is responsible for all four activities. 5. Two forms of the enzyme were found to occur either separately or together in kidneys of pigs from several different breeds. 6. Electro-focusing experiments show these to have a small difference in isoelectric point (4.9 and 5.1), and gel filtration gives an approximate molecular weight of 50000 for both forms. 7. The characteristics of these two enzymes are compared.     

Influence of phospholipid asymmetry on fusion between large unilamellar vesicles.

The ability of lipid asymmetry to regulate Ca(2+)-stimulated fusion between large unilamellar vesicles has been investigated. It is shown that for 100-nm-diameter LUVs composed of dioleoylphosphatidylcholine, dioleoylphosphatidylethanolamine, phosphatidylinositol, and dioleoylphosphatidic acid (DOPC/DOPE/PI/DOPA; 25:60:5:10) rapid and essentially complete fusion is observed by fluorescent resonance energy transfer techniques when Ca2+ (8 mM) is added. Alternatively, for LUVs with the same lipid composition but when DOPA was sequestered to the inner monolayer by incubation in the presence of a pH gradient (interior basic), little or no fusion is observed on addition of Ca2+. It is shown that the extent of Ca(2+)-induced fusion correlates with the amount of exterior DOPA. Further, it is shown that LUVs containing only 2.5 mol % DOPA, but where all the DOPA is in the outer monolayer, can be induced to fuse to the same extent and with the same rate as LUVs containing 5 mol % DOPA. These results strongly support a regulatory role for lipid asymmetry in membrane fusion and indicate that the fusogenic tendencies of lipid bilayers are largely determined by the properties of the monolayers proximate to the fusion interface.     

[2-(3-Nitrotyrosine)]oxytocin, a weakly active analogue of the hormone bound by neurophysin (Short Communication)

An analogue of oxytocin containing a nitro group ortho to the phenolic hydroxyl group of the tyrosyl residue was prepared by nitration of the hormone with tetranitromethane. [2-(3-Nitro-l-tyrosine)]oxytocin was bound by neurophysin although its pharmacological activity was virtually abolished. The oxytocic activity of the analogue on the isolated rat uterus was 1.1i.u./mg.     

Acid-phosphatase activity in sieve-tube members of Tilia americana

Summary Acid phosphatases are localized on the internal strands of sieve-tube members in the secondary phloem of Tilia americana. Companion cells and certain strand parenchyma cells associated with sieve-tube members show a high activity of acid phosphatases.This research has been supported by NSF Grant GB-615.