Galactosyl transferases of baby hamster kidney (BHK) cells. Characterization of two oligosaccharide products synthesised using bovine asialo submaxillary-gland mucin as acceptor

Extracts of BHK (baby hamster kidney) cells catalyse incorporation of galactose from UDP-galactose into asialo bovine submaxillary gland mucin. The galactosylated oligosaccharide products were released by alkaline-borohydride treatment and purified by Bio-Gel P2 chromatography and high-performance liquid chromatography. The structures of the oligosaccharide sequences synthesised have been identified unequivocally by high resolution 500 MHz 1H-NMR as galactosyl-(beta 1----3) N-acetylgalactosamine and galactosyl (beta 1----4) N-acetylglucosaminyl (beta 1----3)-N-acetylgalactosamine. Characterization of the latter sequence shows the presence in bovine mucin of the type III core sequence N-acetylglucosamine-(beta 1----3) N-acetylgalactosamine. Fractionation of BHK cell extracts on alpha-lactalbumin-Agarose has shown that the (beta 1----4)-galactosyl transferase responsible for synthesis of the trisaccharide binds to alpha-lactalbumin, a modulator of the (beta 1----4)-galactosyl transferase involved in N-glycan assembly. The evidence that the same transferase activity may be responsible for galactose transfer to both O-glycans and N-glycans is discussed.     

A simple liquid medium for chlamydospore formation in Candida albicans

A new, relatively simple and inexpensive liquid medium was devised to produce all structural forms ofC. albicans. Optimum conditions to induce the yeast cells, germ tubes, pseudohyphae and chlamydospores along with the methods to obtain them are described.Supported in part by Grant CA 20917, National Cancer Institute, National Institutes of Health and ALSAC.     

Scanning electron microscopy of the respiratory surfaces of Saccobranchus (=Heteropneustes) fossilis (Bloch)

Summary The gill secondary lamellae are generally covered with epithelial cells whose outer surfaces form numerous microvilli. The surface of the primary lamellae is characterised by microridges. A particular type of surface sculpturing seems to be associated with given cell boundaries.Further evidence for the derivation of the air tube and fans which guard its entrance by modification of the basic gill structure has been obtained from both the gross surface architecture and microstructure of the individual cell surfaces. Secondary lamellae are represented by stubby projections which generally have a biserial arrangement. The outer surfaces of the epithelia overlying the capillaries of these respiratory islets are coated with microvilli as in the secondary lamellae. On the other hand, the relatively smooth-surfaced lanes between groups of respiratory islets have a microridged surface similar to that of the primary gill lamellae.It is suggested that previous estimates of surface area, and consequently diffusing capacities of the air-breathing organ, have been low in view of the increased surface, due to both their gross and microstructure. Estimates for gill surface area may need very little correction as the spaces between the microvilli and microridges are probably filled with mucus under normal conditions.We thank Mr. John Clements for his excellent technical assistance and the Department of Botany, Bristol University for the use of their scanning electron microscope     

Mastalgia: psychoneurosis or organic disease?

To test the traditional surgical view that pain in the breast is largely an expression of psychoneurosis, the Middelesex Hospital Questionnaire was given to 317 women with mastalgia and 170 controls with varicose veins. Their scores were compared with those of 173 women psychiatric outpatients tested by the designers of the questionnaire. The results were broadly similar in the mastalgia and varicose veins groups, and where there were significant differences women with varicose veins had a higher psychoneurotic score in each case. Within the mastalgia group no difference in scores was observed between patients with cyclical mastalgia and those with mastalgia due to periductal mastitis. Both groups of surgical outpatients had significantly lower scores in major traits than the psychiatric group, except for a small group of patients with breast pain who persistently failed to respond to treatment. Patients with mastalgia are therefore no more "neurotic" than those with varicose veins, and differ greatly from patients with recognized psychoneurosis. Most patients have a physiological or pathological basis for their breast pain, and they deserve an appropriate diagnostic and therapeutic approach.     

Effects of neuraminidase on lectin binding by wild-type and ricin-resistant strains of hamster fibroblasts.

The nature of cell surface receptors for ricin on wild-type and ricin-resistant variants of baby hamster kidney fibroblasts has been studied. Neuraminidase stimulated ricin binding threefold by wild-type cells, and increased their susceptibility to ricin toxicity as measured by inhibition of [3H]leucine uptake (LD30 fell from 5.0 to 0.5 microgram/mL). Basal ricin binding by ricin-resistant variants (10-300% that of wild type) was also stimulated (2- to 17-fold) by neuraminidase in all seven clonal strains examined; susceptibility to ricin was greatly increased by neuraminidase in these variants. Neuraminidase did not affect the binding of concanavalin A by wild type or a ricin-resistant variant, but decreased the binding of wheat-germ agglutinin by 90% in both cell types. The trivial binding of peanut agglutinin by wild type and a ricin-resistant variant was markedly enhanced (14- to 22-fold) by neuraminidase. Neither collagenase (50 U/mL) nor Pronase (0.0001%) affected ricin binding by wild type or a ricin-resistant variant. These data suggest the existence of "exposed" and "cryptic" oligosaccharide receptors for ricin on the cell membrane glycoproteins of baby hamster kidney fibroblasts. The cryptic ricin receptors probably include at least the sequence D-galactosyl-beta-(1 replaced by 3)-N-acetylhexosamine substituted by sialic acid residues. Exposed and cryptic ricin receptors appear to be different and under separate genetic control.     

An in situ assay for induced diphtheria-toxin-resistant mutants of diploid human fibroblasts

The sensitivity of diploid human fibroblasts to the cytotoxic effects of diphtheria toxin (DT) depended on the cell growth status. Exponentially growing cells treated with 10^?3-1 lethal flocculating units (LF) of DT/ml for 4 days survived with a frequency of 4 × 10^?4. However, the DT-resistant phenotype of colonies isolated under these conditions was not stable. When the growth of the cells had been arrested by confluence or deprivation of serum growth factors prior to treatment with DT (4 days, 10^?3-0.6 LF/ml), the survival decreased to 2 × 10^?6 and the resistance of isolated colonies was stable. An in situ assay for induced DT-resistant mutants was developed in order to avoid problems associated with the possible reduced viability of the mutants relative to that of wild-type cells. A reproducible and linear dose response was obtained for the induction of DT-resistant mutants by ethylnitrosourea. The mutants were induced with high frequency by this compound (e.g., 10^?3 mutants/viable cell at a 37% survival dose); complete expression of the mutant phenotype occurred after 6 generations of growth under nonselective conditions. Isolated mutant colonies showed stable resistance to DT and were cross-resistant to Pseudomonas aeruginosa exotoxin A.     

Diastereoisomers of cyclocymopol and cyclocymopol monomethyl ether from Cymopolia barbata

Optically active diastercoisomers of the brominated monoterpene quinols, cyclocymopol and cyclocymopol monomethyl ether, were isolated from the green marine alga Cymopolia barbata and charac- terized.     

Peptide fragmentation suitable for solid-phase microsequencing. Use of N-bromosuccinimide and BNPS-skatole (3-bromo-3-methyl-2-[(2-nitrophenyl)thio]-3H-indole).

A new intracellular beta-glucosidase was isolated from Trichoderma reesei. It was sequentially purified by (NH4)2SO4 precipitation and chromatography and rechromatography on Sephadex G-150. The enzyme has a mol.wt. of 98 000, optimal activity at pH 6.5, pI 4.4 and Km values of 6.7 mM and 3.3 mM for sophorose and cellobiose respectively. Possible functions of the enzyme may be regulation of cellulase induction and/or to serve as a proenzyme.     

Nearest-neighbor and next-nearest-neighbor effects in the proton NMR spectra of the oligoribonucleotides ApGpX and CpApX

The variable-temperature proton nmr spectra of the oligoribonucleotides in the series CpApX and the series ApGpX, X = A, G, C, U, together with the parent dimers CpA and ApG have been measured. A complete analysis of all the nonexchangeable base proton resonances and ribose H-1′ proton resonances was made. The presence of trends in the shielding abilities of the various bases at both the nearest-neighbor and next-nearest-neighbor positions were identified. The observed shieldings could be used to predict the chemical shifts of protons in related systems. Based on the empirical results from ribodinucleoside monophosphates, the temperature-dependent behavior of the J_1′2′ coupling constants of the triribonucleotides suggested that the compounds in the CpApX series stacked from the 5′-end to the 3′-end, while those in the ApGpX series stacked from the 3′-end to the 5′-end.