Interaction of chlorpromazine with milk proteins

The mode and nature of the binding of chlorpromazine (CPZ), a psychotropic drug, with milk proteins – -lactalbumin (with substantial amounts of -helix, -sheet and random coil), -lactoglobulin (a major -sheeted protein) and _s-casein (a random coiled protein) have been studied spectrofluorometrically and spectropolarimetrically. The binding affinity of CPZ for unfolded proteins is comparatively less than that of folded proteins although the number of binding sites is smaller in the latter case, due to the greater extent of binding of CPZ for folded proteins. Thermodynamic analysis reveals that CPZ binds to -lactalbumin and _s-casein in an endothermic (H^o is positive) and hydrophobic manner but with -lactoglobulin in an exothermic (H^o is negative) manner. Far UV Circular dichroic studies reveal that CPZ increases the secondary structure of the major -sheeted protein, -lactoglobulin possibly by increasing the relative contact orders (non-local contacts) within the residues. On the other hand, for proteins possessing random coil, it increases the unfolded state of the protein. CPZ does not affect local contacts in a-helix when its interaction is compared with a major -helical protein, myoglobin.     

Synthesis of unique proteins at the onset of carbon starvation inEscherichia coli

Summary Escherichia coli bulk protein synthesis continued during the first 3–4 h of carbon starvation at 50–75% that of non-starved (growing) cells. Two-dimensional gel electrophoresis analysis of in vivo pulse-labelled proteins resolved at least 30 polypeptides with new or increased synthesis, relative to total protein synthesis, during this time. Among these polypeptides were several that were also synthesized by ethanol-treatedE. coli (heat-shock proteins). In addition, a number of unique polypeptides were synthesized by carbon-starved cells. These starvation proteins may be involved in survival of the starving bacteria.     

Occurrence of pppApp-synthesizing activity in actinomycetes and isolation of purine nucleotide pyrophosphotransferase

The occurrence of adenosine 5-triphosphate-3-diphosphate-synthesizing activity was detected in five strains of actinomycetes; Streptomyces morookaensis, Streptomyces aspergilloides, Streptomyces hachijoensis, Actinomyces violascens and Streptoverticillium septatum, out of 825 strains of actinomycetes, bacteria, fungi and imperfecti. Purine nucleotide pyrophosphotransferase were extracellularly excreted associating with the cell growth, and were purified partially or to apparent homogeniety from the culture filtrate. The enzymes are a monomeric protein with a molecular weight of 18000–26000 and synthesize adenosine, guanosine and inosine 5-phosphate (mono, di or tri)-3-diphosphate such as pApp, ppApp, pppApp, pGpp, ppGpp, pppGpp and pppIpp by transferring a pyrophosphoryl group from the 5-position of ATP, dATP and pppApp to the 3-position of purine nucleotides in the presence of a divalent cation and in alkaline state.Abbreviations pppApp adenosine 5-triphosphate 3-diphosphate - ppApp adenosine 5-diphosphate 3-diphosphate - pApp adenosine 5-monophosphate 3-diphosphate - pppGpp guanosine 5-triphosphate 3-diphosphate     

The catalysis of nucleotide polymerization by compounds of divalent lead

Summary Soluble lead salts and a number of lead-containing minerals catalyze the formation of oligonucleotides from nucleoside 5-phosphorimidazolides. The effectiveness of lead compounds correlates strongly with their solubility. Under optimal conditions we were able to obtain 18% of pentamer and higher oligomers from ImpA. Reactions involving ImpU gave smaller yields.Abbreviations A adenosine - U uridine - Im imidazole - MeIm 1-methyl-imidazole - EDTA ethylenediaminetetraacetic acid - pA adenosine 5-phosphate - pU uridine 5-phosphate - Ap adenosine cyclic 2:3-phosphate - ATP adenosine 5-triphosphate - AppA P₁,P₂-diadenosine 5-diphosphate - pNp (N = A,U) nucleotide 2(3), 5-diphosphate - ImpA adenosine 5-phosphoreimidazolide - ImpU uridine 5-phosphorimidazolide - A ²pA adenylyl-[25]-adenosine - A ³pA adenylyl-[35]-adenosine - pA ²pA 5-phospho-adenylyl-[25]-adenosine - pA ³pA 5-phospho-adenylyl-[35]-adenosine - pUpU 5-phospho-uridylyl-uridine - pApU 5-phospho-adenylyl-uridine - pUpA 5-phospho-uridylyladenine - (pA)n (n, 2,3,4,) oligoadenylates with 5 terminal phosphate - ImpApA 5-phosphorimidazolide of adenylyl adenosine - (pA) ₅₊ pentamer and higher oligoadenylates with 5 terminal phosphate - (Ap)nA (n = 2,3,4) oligoadenylates without terminal phosphates In the following we do not specify the nature of the internucleotide linkageIn the following we do not specify the nature of the internucleotide linkage     

Puromycin enhances 20-hydroxyecdysone-induced protein synthesis in wing discs of Drosophila melanogaster

The effect of cycloheximide and puromycin on 20-hydroxyecdysone-induced protein synthesis in wing discs of Drosophila melanogaster has been studied by one-dimensional and two-dimensional SDS polyacrylamide electrophoresis. It is found that puromycin, but not cycloheximide, when applied simultaneously with the hormone enhanced the hormone-induced synthesis of the early and late proteins. However, when puromycin was applied after hormone treatment, only the late proteins were induced. The possible implication of these observations is discussed.     

Protonmotive force driven 6-deoxyglucose uptake by the oral pathogen,Streptococcus mutans Ingbritt

Streptococcus mutans Ingbritt was grown in glucose-excess continuous culture to repress the glucose phosphoenolpyruvate phosphotransferase system (PTS) and allow investigation of the alternative glucose process using the non-PTS substrate, (³H) 6-deoxyglucose. After correcting for non-specific adsorption to inactivated cells, the radiolabelled glucose analogue was found to be concentrated approximately 4.3-fold intracellularly by bacteria incubated in 100 mM Tris-citrate buffer, pH 7.0. Mercaptoethanol or KCl enhanced 6-deoxyglucose uptake, enabling it to be concentrated internally by at least 8-fold, but NaCl was inhibitory to its transport. Initial uptake was antagonised by glucose but not 2-deoxyglucose. Evidence that 6-deoxyglucose transport was driven by protonmotive force (p) was obtained by inhibiting its uptake with the protonophores, 2,4-dinitrophenol, carbonylcyanide m-chlorophenylhydrazine, gramicidin and nigericin, and the electrical potential difference () dissipator, KSCN. The membrane ATPase inhibitor, N,N¹-dicyclohexyl carbodiimide, also reduced 6-deoxyglucose uptake as did 100 mM lactate. In combination, these two inhibitors completely abolished 6-deoxyglucose transport. This suggests that the driving force for 6-deoxyglucose uptake is electrogenic, involving both the transmembrane pH gradient (pH) and . ATP hydrolysis, catalysed by the ATPase, and lactate excretion might be important contributors to pH.Abbreviations DNP 2,4-dinitrophenol - CCCP carbonylcyanide m-chlorophenylhydrazone - DCCD N,N¹-dicyclohyxyl carbodiimide - p protonmotive force - pH transmembrane pH gradient - transmembrane electrical potential difference     

Synthesis ofNeoglycoproteins containing the 3,6-di-O-methyl-β-d-glucopyranosyl epitope and their use in serodiagnosis of leprosy

A stratagem for the synthesis ofneoglycoproteins suitable for the selective serodiagnosis of leprosy is described in which synthetic 3,6-di-O-methyl--d-glucopyranose, the epitope of phenolic glycolipid I fromMycobacterium leprae, was used. Condensation of 8-methoxycarbonyloctanol with the acetobromo derivative of 3,6-di-O-methylglucose gave 8-methoxycarbonyloctyl 2,4-di-O-acetyl-3,6-di-O-methyl--d-glucopyranoside in 65% yield, and with absolute stereospecificity for the anomer. The deacylated product was converted to the crystalline hydrazide and coupled to bovine gamma globulin, bovine serum albumin and poly-d-lysinevia intermediate acyl azide formation to produce the 8-carbonyloctyl 3,6-di-O-methyl--d-glucopyranosyl polypeptides. Theneoglycoproteins were highly sensitive in ELISA and emulated the specificity of the native glycolipid in analysis of sera from patients throughout the spectrum of leprosy and from different geographical regions. The 8-carbonyloctyl 3,6-di-O-methyl--d-glucopyranoside-bovine serum albumin was also synthesized and shown to have about one-half the activity of the -linkedneoglycoprotein. A different synthetic approach produced the 8-carbonyloctyl 4-O-(3,6-di-O-methyl--d-glucopyranosyl)--l-rhamnopyranoside-bovine serum albumin which was also highly sensitive and specific for the serodiagnosis of leprosy. The presence of the second sugar unit, similar to that in the native glycolipid but for the absence ofO-methyl groups, seemed to provide a probe with greater felicity for the serological detection of tuberculoid leprosy.Thus, the results indicate that highly sensitive and specific antigen probes for the serodiagnosis of leprosy can be constructed based only on the terminal one or two sugars of phenolic glycolipid I, and the synthetic approach leads to the formation of haptens with absolute stereospecificity.Nomenclature BGG bovine gamma globulin - PGL-I phenolic glycolipid I - PDL poly-d-lysine - PBS phophate-buffered saline - 3,6-Me₂-Glc-Link-BSA 8-carbonyloctyl 3,6-di-O-methyl-glucopyranoside-bovine senalbumin - 3,6-Me₂-Glc-Rha-Link-BSA 8-carbonyloctyl 4-O-(3,6-di-O-methyl--d-glucopyranosyl)--l-rhan pyranoside-BSA     

Comparative effects of sorbitol and sucrose as main carbon energy sources in micropropagation of apricot

In vitro proliferation and rooting capacity of San Castrese and Portici apricots (Prunus armeniaca L.) were tested on modified MS medium enriched with varying growth regulator concentrations and sucrose (58.4 mM) or sorbitol (116.8 mM) as main carbon energy sources. The interaction of proliferation and rooting media was also studied.Proliferation of both cultivars was proportional to benzyladenine (BA) concentration and enhanced with sorbitol media. However, 8.8 M BA was often associated with hyperhydricity, particularly when shoots were grown on sucrose media. Newly proliferated shoots elongated better on sorbitol media. The positive influence of sorbitol on proliferation and shoot growth was not due to osmotic effects. Moreover, sorbitol showed a positive carryover effect in hastening rooting of Portici. By contrast, when transferred to sorbitol rooting media, the shoots of both cultivars generally showed low rooting, with short, thick roots.Up to 70% of the plantlets that produced roots in sucrose media enriched with indolebutyric acid were successfully acclimatized when they were dipped in a benomyl (0.075% w/v) suspension before being transplanted with care being taken to prevent over-wetting of soil.Abbreviations BA 6-benzyladenine - IBA indolebutyric acid - GA₃ gibberellic acid - SEM standard error of mean     

Epigenetic Processes, when Natura Non Facit Saltum Becomes a Myth, and Alternative Ontogenies a Mechanism of Evolution

When I started the journal Environmental Biology of Fishes more than 25 years ago, it was designed to avoid any prejudices and scientific dogmas, to keep an open mind, and to allow publication of ideas that are irritating to followers of the so-called mainstream, ideas that few other journals dared to touch (Balon 1976, 1989d). While such articles in the journal were often ignored by the mainstream practitioners and the political establishment, none was ever opposed or proven wrong. As in many similar cases before, only time will show if these contributions were valid and of value to the advancement of knowledge.I have often wondered why it takes so long for some of the universal facts and ideas to be accepted or even to be known. I have come to believe that like with many textbooks some facts and conclusions are copied from one book to the next (as in the case of the origin of the common carp – Balon 1974, 1995a,b) without knowing or admitting that in the meantime new facts require new syntheses and conclusions. Even good scholars resist unknown facts or ideas. It is irritating that one's knowledge is inadequate or that others beat us to be first. I have wondered why wrong ideas persist for so long; but Bateson (1979, p. 206) already explained it to his daughter ... yes, your image of evolution is exact. And what Darwin called natural selection is the surfacing of the tautology or presupposition that what stays true longer does indeed stay true longer than what stays true not so long.After all it was Jean Baptiste Lamarckgrave, the founder of evolutionary theory, miserable, old, and blind, ... (Bateson 1972, p. xii) and/or Gottfried Treviranus who presented the truth long before the privileged Charles Darwin, alas in wrong languages. Over time, ignoring the truth became fashionable and the question which of the four theories of evolution (Løvtrup 1982) can be attributed to Darwin, if any, irrelevant. It is [exclaims Croizat 1977, transl. 1987, p. 137] a gross, very grave mistake to confuse Darwin as a historical figure with Darwin as an exalted figure of the thinking of biology (Croizat's italics).Nearly 30 years ago a BBC TV series and book on evolution glorified Darwin and molecular biology. Calder (1973, p. 9), the author of this book entitled The life game: evolution and the new biology felt that Darwin would [merely] need to know about the survival of the quickest, when it came to crossing the street. But the street was never crossed! Many TV viewers of the recent PBS series Evolution were again misled into believing that evolution was discovered by Darwin. Even Gould (2001) in the Introduction and the producer Hutton (2001) in the Foreword to the companion book of this series (Zimmer 2001) give the Darwinian evolution [sic] an exposé it hardly deserves. While such views have been repeatedly proven wrong, most scholars, with the exception of a few scientists, philosophers, and journalists, comfortably assume Darwin's priority in spite of numerous published evidences to the contrary. This myth is then fed to the general public. The following retrospective essay attempts to compile proofs in support of the evolutionary processes other than Darwin's natural selection and the gene-centric new synthesis, old by now as it is.     

Spectral characteristics of phytochrome in vivo and in vitro

The absorption maximum of the far-red absorbing form of phytochrome in the difference spectrum for phototransformation (Pfr max) was investigated in vivo and in in vitro pellets from dark grown Hordeum vulgare L. primary leaves. Exposure of pellets in Honda medium from tissue pre-irradiated with red light to far red light gave a Pfr max of 734 nm, a slightly longer wavelength than was seen in vivo (730 nm). After incubation as the red absorbing form of phytochrome (Pr) for 2 h at 0° C irradiation with red light showed that Pfr max had shifted to shorter wavelength (716 nm) in Honda medium. Further incubation as Pfr for 2 h at 0° C and irradiation with far red light showed that Pfr max had shifted to longer wavelength (726 nm). Similar shifts were also seen in other media, although the peak positions were different. Phytochrome remained pelletable throughout these experiments and Pfr max is compared to that of soluble phytochrome in similar media. The results are interpreted as indicating changes in molecular environment of the putative phytochrome membrane receptor site and that Pfr max can be used to probe the nature of this binding.Abbreviations D Dark - EDTA Ethylene diamine tetra-acetic acid - F far red light - MOPS N-morpholino-3-propane-sulphonic acid - P Phytochrome - Pr red absorbing form of P - Pfr far red absorbing form of P - Pfr max wavelength maximum of Pfr absorbance in a phototransformation difference spectrum - R red light     

Comparison of the homologous carboxy-terminal domain and tail of α-crystallin and small heat shock protein

The C-terminal domain and tail, which is the most conserved region of the -crystallin/small heat shock protein (HSP) family, was obtained from rat A-crystallin, bovine B-crystallin and mouse HSP25. All three domains have primarily -sheet conformation and less than 10% of -helix, like the proteins from which they are derived. Whereas the C-terminal part of A-crystallin forms dimers or tetramers, the corresponding regions of B-crystallin and HSP25 form larger aggregates. The heat-protective activity, recently described for the -crystallin/small HSP family, is not retained in the C-terminal domain and tail. In the course of this study some differences with the previously published sequence of HSP25 were observed, and a revision is proposed.Abbreviations A2Dt residues 64–173 of rat -crystallin - B2Dt residues 70–175 of bovine B-crystallin - bp base pair - HSP2Dt residues 92–209 of HSP25 - HSP(s) heat shock protein(s) - HSP25 mouse small HSP - PCR polymerase chain reaction - PMSF phenylmethylsulfonyl chloride - SDS sodium dodecyl sulfate; polyacrylamide - WSF water-soluble fraction     

Asparagine metabolism and nitrogen distribution during protein degradation in sugar-starved maize root tips

Excised maize (Zea mays L.) root tips were used to monitor the effects of prolonged glucose starvation on nitrogen metabolism. Following root-tip excision, sugar content was rapidly exhausted, and protein content declined to 40 and 8% of its initial value after 96 and 192 h, respectively. During starvation the contents of free amino acids changed. Amino acids that belonged to the same synthetic family showed a similar pattern of changes, indicating that their content, during starvation, is controlled mainly at the level of their common biosynthetic steps. Asparagine, which is a good marker of protein and amino-acid degradation under stress conditions, accumulated considerably until 45 h of starvation and accounted for 50% of the nitrogen released by protein degradation at that time. After 45 h of starvation, nitrogen ceased to be stored in asparagine and was excreted from the cell, first as ammonia until 90–100 h and then, when starvation had become irreversible, as amino acids and aminated compounds. The study of asparagine metabolism and nitrogen-assimilation pathways throughout starvation showed that: (i) asparagine synthesis occurred via asparagine synthetase (EC 6.3.1.1) rather than asparagine aminotransferase (EC 2.6.1.14) or the -cyanoalanine pathway, and asparagine degradation occurred via asparaginase (EC 3.5.1.1); and (ii) the enzymic activities related to nitrogen reduction and assimilation and amino-acid synthesis decreased continuously, whereas glutamate dehydrogenase (EC 1.4.1.2–4) activities increased during the reversible period of starvation. Considered together, metabolite analysis and enzymic-activity measurements showed that starvation may be divided into three phases: (i) the acclimation phase (0 to 30–35 h) in which the root tips adapt to transient sugar deprivation and partly store the nitrogen released by protein degradation, (ii) the survival phase (30–35 to 90–100 h) in which the root tips expel the nitrogen released by protein degradation and starvation may be reversed by sugar addition and (iii) the cell-disorganization phase (beyond 100 h) in which all metabolites and enzymic activities decrease and the root tips die.Abbreviations AlaAT alanine aminotransferase - AspAT aspartate aminotransferase - AS asparagine synthetase - Asnase asparaginase - AsnAT asparagine aminotransferase - -CS -cyanoalanine synthase - GDH glutamate dehydrogenase - Glnase glutaminase - GOGAT glutamate synthase - GS glutamine synthetase - NiR nitrite reductase - NR nitrate reductase     

Early generation intermating for yield improvement in groundnut (Arachis hypogaea L.)

Summary Seeds of 4 crosses of groundnut (Arachis hypogaea L.), Robut 33-1 x Chico, Robut 33-1 x NC Ac 17090, Robut 33-1 x PI 298115 and MK 374 x GAUG 1, were irradiated with 30 kR. In the F₁, some branches of each plant were intermated with other plants at random and others selfed in each cross to produce S₂ and F₂ seeds. They were evaluated for pod yield, shelling percentage and 100-kernel weight. The frequency of plants superior to F[in1] was much higher in S₂ than in F₂, which was, in general, true for the values of yield and its components. The S₂ and F₂ were advanced to third generation by selfing. The families descending from S₂ showed clear superiority over those from F₂. The reason for such superiority was suggested to be the recombination of genes from the upper and lower ends of the genotypic distribution under intermating.     

Role of guanosine tetraphosphate in gene expression and the survival of glucose or seryl-tRNA starved cells of Escherichia coli K12

The concentration of guanosine 3,5-bispyrophosphate (ppGpp) increases in bacteria in response to amino acid or carbon/energy source starvation. An Escherichia coli K12 relAspoT mutant lacking the ability to synthesize ppGpp lost viability at an increased rate during both glucose and seryl-tRNA starvation. Also, the deleterious effect of chloramphenicol on starved wild-type cells could be overcome by inducing expression of RelA from a plasmid carrying the relA gene transcribed from a tac promoter, prior to starvation and chloramphenicol treatment. As demonstrated by two dimensional gel electrophoresis, this induction of the RelA protein resulted in global alterations in gene expression including increased synthesis of some rpoS-dependent proteins. The relAspoT mutant maintained high expression of several ribosomal proteins during starvation and appeared to exhibit significantly decreased translational fidelity, as demonstrated by an unusual heterogeneity in the isoelectric point of several proteins and the failure to express higher molecular weight proteins during starvation. Moreover, both rpoS-dependent and independent genes failed to exhibit increased expression in the mutant. It is suggested that the deleterious effects on the cells of the relA, spoT deletions are not due solely to the inability of these cells to induce the sigma factor ^s, but also to deficiencies in translational fidelity and failure to exert classical stringent regulation.     

Substitutions of the Conserved Thr42 Increased the Roles of the ε-Subunit of Maize CF1 as CF1 Inhibitor and Proton Gate

The conserved residue Thr42 of -subunit of the chloroplast ATP synthase of maize (Zea mays L.) was substituted with Cys, Arg, and Ile, respectively, through site-directed mutagenesis. The over-expressed and refolded -proteins were purified by chromatography on DEAE-cellulose and FPLC on mono-Q column, which were as biologically active (inhibiting Ca²⁺-ATPase activity and blocking proton gate) as the native subunit isolated from chloroplasts. The T42C and T42R showed higher inhibitory activities on the soluble CF₁(–) Ca²⁺-ATPase than the WT. The T42I inhibited the Ca²⁺-ATPase activity of soluble CF₁ and restored photophosphorylation activity of membrane-bound CF₁ deficient in the most efficiently. Far-ultraviolet CD spectra showed that the portions of -helix and -sheet structures of the three mutants were somewhat different from WT. Thus the conserved residue Thr42 may be important for maintaining the structure and function of the -subunit and the basic functions of the -subunit as far as an inhibitor of Ca²⁺-ATPase and the proton gate are related.     

The in vitro physiological phenotype of tomato resistance to Fusarium oxysporum f. sp. lycopersici

Summary With the aim of dissecting host-parasite interaction processes in the system Lycopersicon aesculentum-Fusarium oxysporum f. sp. lycopersici we have isolated plant cell mutants having single-step alterations in their defense response. A previous analysis of the physiological phenotypes of mutant cell clones suggested that recognition is the crucial event for active defence, and that polysaccharide content, fungal growth inhibition, peroxidase induction in in vitro dual culture and ion leakage induced by cultural filtrates of the pathogen can be markers of resistance. In this paper we present the results of a similar analysis carried out on cell cultures from one susceptible (Red River), one tolerant (UC 105) and three resistant (Davis UC 82, Heinz, UC 90) tomato cultivars. Our data confirm that the differences in the parameters considered are correlated with resistance versus susceptibility in vivo. Therefore, these parameters can be used for early screening in selection programmes. These data, together with those obtained on isolated cell mutants, suggest that the selection in vitro for altered fungal recognition and/or polysaccharide or callose content may lead to in vivo — resistant genotypes. The data are thoroughly discussed with particular attention paid to the importance of polysaccharides in active defense initiation.     

Effect of starvation on the content of free amino acids in plasma of different breeds of hen

Summary Leghorn, Cornish and White Rock hens were subjected to starvation. Free amino acids were determined in blood samples taken after 48, 72 and 96 h of starvation. A progressive decrease in concentration of the majority of amino acids was found. Changes in amino acid concentrations during starvation were dependent on the breed of hen.     

Inheritance of mitochondrial DNA in lentil (Lens culinaris Medik.)

Restriction fragment analysis was used to examine the inheritance of lentil mitochondrial DNA (mtDNA) in F₁ and F₅ progeny from intrasubspecific (Lens culinaris ssp. culinaris) crosses and in F₁ progeny from intersubspecific (Lens culinaris ssp. orientalis x L. culinaris ssp. culinaris) crosses. Southern blots of digested parental and progeny DNA were hybridized to heterologous maize mtDNA probes specific to coxI and atp6 genes. Two restriction fragment polymorphisms separated L.c. ssp. culinaris Laird and Eston from L.c. ssp. culinaris ILL5588, and one restriction fragment polymorphism distinguished L.c. ssp. culinaris Laird and Eston from L.c. ssp. orientalis LO4. Twelve of 13 f₁ progeny and all F₅ progeny from the intrasubspecific crosses, and all F₁ progeny from intersubspecific crosses had only maternal mtDNA restriction fragments. One f₁ plant from an Eston x ILL5588 cross inherited mtDNA fragments from both parents. Nuclear DNA inheritance was biparental in all F₁ progeny.NRCC No. 38451     

Sialyltransferase activity in FR3T3 cells transformed withras oncogene: decreased CMP-Neu5Ac:Galβ1-3GalNAc α-2,3-sialyltransferase

We have investigated the activity of CMP-Neu5Ac:Gal\1-3GalNAc -2,3-sialyltransferase (EC 2.4.99.4) in FR3T3 cells transformed by the Ha-ras oncogene in which we have previously demonstrated the higher expression of the -galactosidase -2,6-sialyltransferase (EC 2.4.99.1) [21]. We demonstrate that the presence of the activatedras gene decreases the activity of this specific -2,3-sialyltransferase fourfold. According to the kinetic parameters and to mixing experiments, we can assume that this decreased enzymatic activity reflects a decrease in the number of activeO-glycan -2,3-sialyltransferase polypeptides inras-transformed cells. However, no change in the binding of Peanut agglutinin was observed on the cell surface ofras-transformed FR3T3 suggesting that no change in the sialylation ofO-glycan core 1 appeared in these cells, although the activity of the -2,3-sialyltransferase was decreased.Abbreviations -2,3-ST(O) CMP-Neu5Ac:Gal1-3GalNAc-R -2,3-sialyltransferase - -2,3-ST(N/O) CMP-Neu5Ac:Gal1-3/4GlcNAc-R -2,3-sialyltransferase - -2,6-ST(N) CMP-Neu5Ac:Gal1-4GlcNAc-R -2,6-sialyltransferase - -2,6-ST(O)I CMP-Neu5Ac:R-GalNAc(1-O)Ser -2,6-sialyltransferase - -2,6-ST(O)II CMP-Neu5Ac:Neu5Ac2-3Gal1-3GalNAc-R -2,6-sialyltransferase - ASFet asialofetuin - FR3T3 Fisher rat fibroblast - FRras Ha-ras-transfected FR3T3 fibroblasts - NaCl/P_i sodium phosphate 10mm, NaCl 0.15m, pH 7.4, buffer - pNp p-nitrophenol     

Nuclear Overhauser effect investigation on GM1 ganglioside containingN-glycolyl-neuraminic acid (II3Neu5GcGgOse4Cer)

The conformational properties of the oligosaccharide chain of GM1 ganglioside containingN-glycolyl-neuraminic acid, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer, were studied through NMR nuclear Overhauser effect investigations on the monomeric ganglioside in dimethylsulfoxide, and on mixed micelles of ganglioside and dodecylphosphocholine in water. Several interresidual contacts for the trisaccharide core--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-were found to fix the relative orientitation of the three saccharides, while the glycosidic linkage of the terminal -Gal-was found to be quite mobile as the -Gal-(1-3)--GalNAc-disaccharide exists in different conformations. These results are similar to those found for two GM1 gangliosides containingN-acetyl-neuraminic acid and neuraminic acid [1].Abbreviations Ganglioside nomenclature is in accordance with Svennerholm [23] and the IUPAC-IUB Recommendations [24] GM3(Neu5Ac) II³Neu5AcLacCer, -Neu5Ac-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM3(Neu5Gc) II³Neu5GcLacCer, -Neu5Gc-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Ac) II³Neu5AcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Gc) II³Neu5GcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu) II³NeuGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu-(2-3)]--Glc-(1-1)-Cer - GD1a IV³Neu5AcII³Neu5AcGgOse₄Cer, -Neu5Ac-(2-3)--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GalNAc-GD1a IV⁴GalNAcIV³Neu5AcII³Neu5AcGgOse₄Cer, -GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - Neu neuraminic acid - Neu5Ac N-acetyl-neuraminic acid - Neu5Gc N-glycolyl-neuraminic acid - Cer ceramide