Enzymic synthesis,chemical characterisation and Sda activity of GalNAcß1-4[NeuAcα2-3]Galß1-4GlcNAc and GalNAcß1-4[NeuAcα2-3]Galß1-4Glc

The tetrasaccharides GalNAcß1-4[NeuAc2-3]Galß1-4Glc and GalNAcß1-4[NeuAc2-3]Galß1-4GlcNAc were synthesised by enzymic transfer of GalNAc from UDP-GalNAc to 3-sialyllactose (NeuAc2-3Galß1-4Glc) and 3-sialyl-N-acetyllactosamine (NeuAc2-3Galß1-4GlcNAc). The structures of the products were established by methylation and¹H-500 MHz NMR spectroscopy. In Sd^a serological tests the product formed with 3-sialyl-N-acetyllactosamine was highly active whereas that formed with 3-sialyllactose had only weak activity.     

Effects of etretinate on the distribution of elements in rats

We studied in the rat the effects of the drug etretinate (Tigason), given at three doses 3, 10, and 30 mg/kg body wt for 1 mo, on the concentrations of Na, K, Ca, Mg, Fe, S, P, Cu, and Zn in the plasma, brain, thymus, heart, liver, lung, kidney, testicle, muscle, and bone. The elements were simultaneously determined in tissues after nitric acid dissolution by inductively coupled plasma emission spectrometry using a JY 48 instrument. At the dose of 3 mg/kg, etretinate did not induce any statistically significant modifications of the element distribution. At the dose of 10 mg/kg, the main observed modifications were in plasma an increase of copper (+38%) and a decrease of zinc (-25%). At the highest dose of 30 mg/kg, some variations of the concentrations of elements in tissues were observed. But, on no account did retinoids induce an alteration of the mineral composition of bone, despite obvious macroscopic bone alterations.     

Ouabain-insensitive salt and water movements in duck red cells. II. Norepinephrine stimulation of sodium plus potassium cotransport

Catecholamines induce net salt and water movements in duck red cells incubated in isotonic solutions. The rate of this response is approximately three times greater than a comparable effect observed in 400 mosmol hypertonic solutions in the absence of hormone (W.F. Schmidt and T. J. McManus. 1977 a.J. Gen. Physiol. 70:59-79. Otherwise, these two systems share a great many similarities. In both cases, net water and salt movements have a marked dependence on external cation concentrations, are sensitive to furosemide and insensitive to ouabain, and allow the substitution of rubidium for external potassium. In the presence of ouabain, but the absence of external potassium (or rubidium), a furosemide-sensitive net extrusion of sodium against a large electrochemical gradient can be demonstrated. When norepinephrine-treated cells are incubated with ouabain and sufficient external sodium, the furosemide-sensitive, unidirectional influxes of both sodium and rubidium are half- maximally saturated at similar rubidium concentrations; with saturating external rubidium, the same fluxes are half-maximal at comparable levels of external sodium. In the absence of sodium, a catecholamine-stimulated, furosemide-sensitive influx of rubidium persists. In the absence of rubidium, a similar but smaller component of sodium influx can be seen. We interpret these results in terms of a cotransport model for sodium plus potassium which is activated by hypertonicity or norepinephrine. When either ion is absent from the incubation medium, the system promotes an exchange-diffusion type of movement of the co-ion into the cells. In the absence of external potassium, net movement of potassium out of the cell leads to a coupled extrusion of sodium against its electrochemical gradient.     

Enhanced bismuth digestive absorption in rats by some sulfhydryl compounds: nmr study of complexes formed

In a preliminary paper [Therapie 34, 397 (1979), we showed that cysteine enhances bismuth digestive absorption in rats. In this paper, we have studied in rats the effects of various thiol compounds (mercaptopropionic acid, penicillamine, cysteine, homocysteine, 2-mercaptoethylamine, mercaptoethane) and nonthiol compounds (methionine, serine, alanine) orally administered on the absorption and elimination of bismuth also given orally. Bismuth was measured in blood and urine by electrothermal atomic absorption spectrophotometry.All of the thiol substances, and particularly cysteine, homocysteine, and mercaptopropionic acid, have considerably enhanced bismuth absorption and elimination: whereas, nonthiol substances have had no effect. Moreover, the acute toxicity of bismuth was enhanced when bismuth was given as a complex with cysteine (LD₅₀ = 156± 20 mg/kg).Studies by nmr spectroscopy of interactions between bismuth and these organic compounds have shown that bismuth induces an important chemical shift of the protons of the alpha carbon of the sulfhydrile group. Mainly, studies of ¹³C and ¹⁵N have confirmed this fact. The selectivity of such a complexation, in our pH conditions, may be tentatively explained on the ground of hard and soft acid and base (HSAB) theory.We have suggested that an increase in the concentration of thiol compounds in the gas-trointestinal tract arising from food, or more probably from microorganism synthesis, could be an explanation for human encephalopathies.     

Influence of thyroparathyroidectomy and thyroxine replacement on CU and ZN cellular distribution and on the metallothionein level and induction in rats

Thyroid hormones are involved in copper and zinc distribution in rat tissues. We examined the influence of thyroparathyroidectomy (TPTY) and of a replacement therapy by T4 on Cu and Zn organ distribution. MT levels were also measured both in basal conditions and after induction by cadmium. The results confirm that a lack of T4 modified Cu and Zn in serum and tissues. In serum, TPTY increased Cu (+15%) and ceruloplasmin (+18%), and decreased Zn (−18%). In tissues, Cu was altered in liver (+13%), kidney (−24%), heart (−16%) duodenum (−18%), and Zn in liver (+25%) and kidney (−10%). The soluble fractions (100,000 g supernatant) were mainly affected in liver and kidney, and the subcellular fractions in heart and duodenum. MT levels were modified in basal conditions only in liver (+57%) and kidney (−36%). T4 administration partially prevented the effect of TPTY on both elements and MT concentrations. Therefore, no evidence is provided for a direct role of T4 in the metabolism of MT in a way comparable to the effects of glucocorticoids. However, MT could mediate the consequences of TPTY on metal distribution in certain organs, such as liver and kidney.     

Modifications by BCG of the mortality of the irradiated mouse]

Freeze-dried Bacillus Calmette Guerin (B.C.G.) of Institut Pasteur was given by intravenous route to mice at 1,2 and 4 mg/kg before and after gamma irradiation of animals by 1 000 rad. B.C.G. 1 mg/kg injected the day or the day after irradiation has a protective effect (mortality reduced from 77% for controls to 58% and 50% for treated mice). B.C.G. given before irradiation in single or double doses increased mortality.     

In-depth qualitative and quantitative analysis of composite glycosylation profiles and other micro-heterogeneity on intact monoclonal antibodies by high-resolution native mass spectrometry using a modified Orbitrap

Here, we describe a fast, easy-to-use, and sensitive method to profile in-depth structural micro-heterogeneity, including intricate N-glycosylation profiles, of monoclonal antibodies at the native intact protein level by means of mass spectrometry using a recently introduced modified Orbitrap Exactive Plus mass spectrometer. We demonstrate the versatility of our method to probe structural micro-heterogeneity by describing the analysis of three types of molecules: (1) a non-covalently bound IgG4 hinge deleted full-antibody in equilibrium with its half-antibody, (2) IgG4 mutants exhibiting highly complex glycosylation profiles, and (3) antibody-drug conjugates. Using the modified instrument, we obtain baseline separation and accurate mass determination of all different proteoforms that may be induced, for example, by glycosylation, drug loading and partial peptide backbone-truncation. We show that our method can handle highly complex glycosylation profiles, identifying more than 20 different glycoforms per monoclonal antibody preparation and more than 30 proteoforms on a single highly purified antibody. In analyzing antibody-drug conjugates, our method also easily identifies and quantifies more than 15 structurally different proteoforms that may result from the collective differences in drug loading and glycosylation. The method presented here will aid in the comprehensive analytical and functional characterization of protein micro-heterogeneity, which is crucial for successful development and manufacturing of therapeutic antibodies     

Solution structure of the loop B domain from the hairpin ribozyme

The hairpin ribozyme is a small catalytic RNA with a unique two-domain structure. Here we present the solution structure of the loop B domain of the hairpin ribozyme, which contains most of the catalytically essential nucleotides. The 38-nucleotide domain contains a 16-nucleotide internal loop that forms one of the largest non-Watson-Crick segments of base pairing thus far determined by either NMR or crystallography. Since the solution structure of the smaller loop A domain has been previously solved, an NMR structure-based model of the 22,000 Mr hairpin ribozyme-substrate open complex emerges by joining the two domain structures. Strikingly, catalytically essential functional groups for the loop B domain are concentrated within an expanded minor groove, presenting a clear docking surface for the loop A domain.