Calorimetric and Fourier transform infrared spectroscopic studies on the interaction of glycophorin with phosphatidylserine/dipalmitoylphosphatidylcholine-d62 mixtures

Glycophorin has been isolated in pure form from human erythrocyte membranes and reconstituted into lipid vesicles composed of binary mixtures of bovine brain phosphatidylserine (PS) and acyl-chain perdeuterated dipalmitoylphosphatidylcholine (DPPC-d62). The effect of protein on lipid melting behavior and order has been monitored with differential scanning calorimetry and Fourier transform infrared spectroscopy (FT-IR). The phase diagram for PS/DPPC-d62 is consistent with that previously reported for PS/DPPC (Stewart et al. (1979) Biochim. Biophys. Acta 556, 1-16) and indicates that acyl chain perdeuteration does not greatly alter the lipid mixing characteristics. The use of deuterated lipid allows the examination of lipid order by FT-IR of each lipid component in the binary mixtures as well as in the ternary (lipid/lipid/protein) systems. Addition of glycophorin to a 30:70 PS/DPPC-d62 binary lipid mixture results in a preferential glycophorin/PS interaction leading to bulk lipid enriched in DPPC-d62. This is revealed in two ways: first, through cooperative calorimetric transitions increased in temperature from the binary lipid system and second, through FT-IR melting curves of the DPPC-d62 component which shows transitions increased in both onset and completion temperatures in the presence of protein. In addition, non-cooperative melting events are observed at temperatures below the onset of phase separation. The FT-IR data are used to assign these non-cooperative events to the melting of the PS component. For the 50:50 lipid mixture with protein, two transitions are observed in the DSC experiments. The IR results indicate that both lipid components are involved with the lower temperature event.     

Use of deuterated phospholipids in raman spectroscopic studies of membrane structure. I. Multilayers of dimyristoyl phosphatidylcholine (and its -d54 derivative) with distearoyl phosphatidylcholine

The temperature dependence of the Raman spectrum has been studied for binary phospholipid mixtures of dimyristoyl phosphatidylcholine (and its chain deuterated $\text{-d}{}_{54}$ derivative) with distearoyl phosphatidylcholine. Two distinct melting regions are observed for the 1 : 1 mole ratio mixture. The use of deuterated phospholipid permits the identification of the lower (≈22°C) transition with primarily the melting of the shorter chain component, and the higher (≈47°C) transition primarily with the melting of the longer chains. The C-H stretching vibrations of the distearoyl component respond to the melting of the dimyristoyl component, an apparent consequence of alterations in the lateral interactions of the distearoyl chains. These changes in the C-H spectral region suggest that phase separation does not occur in the gel state for this system. The results are in reasonable accord with recent calorimetric studies (Mabrey, S. and Sturtevant, J.M. (1976) Proc. Natl. Acad. Sci. U.S. 73, 3862–3866). The feasibility of using deuterated phospholipids to monitor the conformation of each component in a binary phospholipid mixture is demonstrated.     

Enzymatic Protein Carboxyl Methylation in Rat Posterior Pituitary: Neurophysins in Rapid-Turnover Pool Determine Methyl Accepting Capacity

In vitro stimulation of intact rat posterior pituitary by either veratridine or K+ depolarization results in the concomitant release of neurophysins and in a decrease (70-80%) in their carboxyl methylation as measured either with L-[methyl-3H]methionine in the intact lobes after stimulation or in their homogenates with [methyl-3H]S-adenosyl-L-methionine and purified protein carboxyl methyltransferase. A similar reduction in neurophysin methylation (60%) was observed when the arrival of newly synthesized neurophysins at the posterior pituitary was blocked by colchicine. Experimental data indicate that the reduction in neurophysin content of the lobes after 12 h of colchicine treatment (less than 7%) or after in vitro stimulation (about 10%) cannot account for the marked reduction in neurophysin methylation. The results suggest that the granule pool characterized by rapid turnover of neurophysins probably represents the major source of methyl acceptor proteins in the lobe. In spite of the marked reduction in neurophysin methyl accepting capacity observed after stimulation, there was no parallel increase in methyl accepting capacity of the released neurophysins. We propose that a neurophysin subfraction that might be associated with the membrane of releasable granules participates in the methylation reaction in situ.     

Liver glutathione S-transferase polymorphism in Japanese and its pharmacogenetic importance

Summary A total of 168 autopsy liver extracts from Japanese individuals were examined for the glutathione S-transferase (GST) isozymes by means of starch gel electrophoresis. The gene frequencies of GST1^*1, GST1^*2, and GST1^*0 in Japanese were 0.252, 0.057, and 0.691, respectively. GST1^*3 was detected as a rare variant allele. The incidence of GST1 0 in 41 liver biopsy samples from patients suffering from various liver diseases was investigated using polyacrylamide gel isoelectric focusing. The GST1 0 phenotype was found more frequently in livers with hepatitis and carcinoma than in control livers. The isozymes coded by different GST loci were partially purified and characterized to study their biochemical properties. The apparent Km values with 1-chloro-2,4-dinitrobenzene (CDNB) as substrate for the isozymes at the GST1, GST2, GST3, and GST4 loci were 604, 1345, 776 and 591 M, respectively.     

Cloning and expression of a functional fucose-specific lectin from an orange peel mushroom, Aleuria aurantia

Aleuria aurantia lectin (AAL) shows sugar-binding specificity for L-fucose. A λgt11 expression library was constructed from A. aurantia poly(A) RNA and screened with a polyclonal antiserum directed against AAL. An immunopositive clone carrying 1.3-kb EcoRI fragment was obtained. The fragment encoded AAL, but lacked a nucleotide sequence corresponding to the two amino-terminal amino acids. The 5′-terminal part of the fragment was replaced with a chemically synthesized DNA fragment and inserted into an expression vector to yield a plasmid pKA-1. Escherichia coli carrying pKA-1 expressed functional AAL and the recombinant AAL showed the same immunological properties as those of natural AAL.     

Local distribution of the lycaenid butterfly,Jalmenus evagoras,in response to host ants and plants

Summary The caterpillars of Jalmenus evagoras are tended by ants as they feed upon Acacia trees. In the area of Brisbane, Australia, J. evagoras require ants of the Iridomyrmex anceps species group; predation and parasitism are so intense that larvae and pupae deprived of attendant ants cannot survive (Pierce 1983). We investigated the efficiency with which J. evagoras locate and exploit the host ant resource by sampling 737 quadrats in 30 sampling grids and six study sites containing appropriate host plants; ants were collected at baits located in the center of each quadrat. J. evagoras was found in all habitats where I. anceps cooccurred with host Acacia. Nine of the ten sampling grids which had three or more I. anceps/Acacia host quadrats also had colonies of J. evagoras present (or immediately adjacent), including sites as far as 35 km apart. Of 19 sampling grids on which host quadrats were rare (i.e., less than three quadrats), none had J. evagoras (P<0.001). Within sample grids, I. anceps was distributed indepedently from Acacia trees, suggesting that they are not dependent for their survival on either Acacia or on J. evagoras. Within montane pasture habitats, I. anceps and at least one other ground-dwelling Iridomyrmex species were distributed in mutually exclusive ant mosaic territories which were stable during a one month period. I. anceps did not colonize or tend pupae of J. evagoras experimentally placed in adjacent territories of a different, nontending species of Iridomyrmex, demonstrating the integrity of territory boundaries. Sampling of ants in Acacia trees revealed that, in the absence of J. evagoras, Iridomyrmex workers are not common above ground level, and that their numbers decline in larger trees (P=0.02). In I. anceps territories, eight of nine J. evagoras pupae placed in trees over 3.0 m tall were not found after 24 h whereas all ten controls placed in low trees were found and tended (P=0.00012). This may explain why J. evagoras tends to oviposit in trees less than 2.0 m tall. An alternative hypothesis, that smaller trees have higher content of total nitrogen, and are threfore more nutritious, was not supported. We conclude that the local distribution and host tree selection by J. evagoras is dependent upon the distribution, patchiness, and foraging behavior of the host ant, I. anceps, and its spatial overlap with a number of species of host Acacia.