Construction of linear GlcNAcβ1-6Galβ1-OR type oligosaccharides by partial cleavage of GlcNAcβ1-3(GlcNAcβ1-6)Galβ1-OR sequences with jack bean β-N-acetylhexosaminidase

Radiolabelled GlcNAc beta 1-3(GlcNAc beta 1-6)Gal (1), GlcNAc beta 1-3)GlcNAc beta 1-6)Gal beta 1-OCH3 (4), GlcNAc beta 1-3(GlcNAc beta 1-6)Gal beta 1-4Glc (7), and GlcNAc beta 1-3(GlcNAc beta 1-6)Gal beta 1-4GlcNAc (10) were cleaved partially with jack bean beta-N-acetylhexosaminidase (EC 3.2.1.30), and the digests were analysed chromatographically. All four oligosaccharides were hydrolysed faster at the (1-6) branch, than at the (1-3) branch, but a high branch specificity was observed only with the glycan 4. The saccharides 1 and 7 resembled each other in the kinetics of the enzyme-catalysed release of their two non-reducing N-acetylglucosamine units, but the glycan 10 was rather different. The partial digestions made it possible to obtain radiolabelled GlcNAc beta 1-6Gal, GlcNAc beta 1-6Gal beta 1-OCH3, GlcNAc beta 1-6Gal beta 1-4Glc, and, in particular, GlcNAc beta 1-6Gal beta 1-4GlcNAc.     

The function of selenocysteine synthase and SELB in the synthesis and incorporation of selenocysteine.

The selAB operon codes for the proteins selenocysteine synthase and SELB which catalyse the synthesis and cotranslational insertion of selenocysteine into protein. This communication deals with the biochemical characterisation of these proteins and in particular with their specific interaction with the selenocysteine-incorporating tRNA(Sec). Selenocysteine synthase catalyses the synthesis of selenocysteyl-tRNA(Sec) from seryl-tRNA(Sec) in a pyridoxal phosphate-dependent reaction mechanism. The enzyme specifically recognizes the tRNA(Sec) molecule; a cooperative interaction between the tRNA binding site and the catalytically active pyridoxal phosphate site is suggested. SELB is an EF-Tu-like protein which specifically complexes selenocysteyl-tRNA(Sec). Interaction with the selenol group of the side chain of the aminoacylated residue is a prerequisite for the formation of a stable SELB.tRNA complex. Mechanistically, this provides the biochemical basis for the exclusive selection of selenocysteyl-tRNA(Sec) in the decoding step of a selenocysteine-specific UGA triplet.     

Penicillium verrucosum in feed of ochratoxin A positive swine herds

Ochratoxin A contamination of cereal feed grain was monitored during October 1989–September 1990 by analysis of blood samples from slaughter swine in Sweden. The detection of ochratoxin A in swine blood was used as a method to identify swine herds fed ochratoxin A contaminated feed. The contamination level of ochratoxin A in the blood of the positive herds was in the range 2–45 ng/ml with the mean concentration 5.2 ng/ml. Feed samples for mycological analysis were collected from both ochratoxin A positive herds (2 ng/ml blood) and ochratoxin A negative herds (<2 ng/ml blood). From the ochratoxin A positive herds and the ochratoxin A negative herds 22 and 21 feed samples were collected, respectively. No quantitative differences in mould content, as determined by colony forming units, were observed between the two groups. However, there were differences in the mycoflora. The incidence of storage fungi (Penicillium and Aspergillus spp.) was significantly higher (p < 0.05) in feed from ochratoxin A positive herds. Particularly, Penicillium verrucosum was found to be significantly more common (p < 0.001). Altogether 274 isolates were screened for their ability to produce ochratoxin A. Ochratoxin A producers were found only within P. verrucosum; 38% of the 63 isolates produced detectable amounts of ochratoxin A. Ochratoxin A producing isolates of P. verrucosum were found in 60% of the feed samples collected from ochratoxin A positive swine herds and in one sample (5% ) of the feed samples collected from the ochratoxin A negative herds.     

Biochemical and spectroscopic analysis of UV effects in the marine flagellate Cryptomonas maculata

The effects of solar and artifical ultraviolet radiation on the marine cryptoflagellate, Cryptomonas maculata, were studied. Even after short exposure to UV the accessory photosynthetic pigment phycoerythrin is bleached; likewise the fluorescence undergoes significant changes both in amplitude and in the maximal peak wavelength. In parallel, the photosynthetic oxygen production decreases rapidly during exposure. Gel electrophoresis and FPLC of membrane proteins show a significant decrease in chromoproteins after 2 h UV, which is confirmed by fluorescence excitation and emission spectra of the FPLC fractions.Abbreviations APS ammonium persulfate - DCMU 3-(3,4dichlorophenyl)1,1-dimethylurea; Emulphogen, polyoxyethylene 10 tridecyl ether - FPLC fast protein liquid chromatography - PMSF phenylmethylsulfonyl fluoride - SDS sodium dodecylsulfate - SDS PAGE sodium dodecylsulfate polyacrylamide gel electrophoresis - TEMED NN NNtetramethylethylene diamine - UV-A wavelength range between 320 nm and 400 nm - UV-B wavelength range between 280 nm and 320 nm Dedicated to the 60th birthday of Professor Dr. W. Wehrmeyer     

Isolation and characterization of a lectin from the cephalochordate Branchiostoma lanceolatum (Pallas).

1. A lectin was isolated from an extract of Branchiostoma lanceolatum by affinity chromatography using an asialo-A-peptone-cellulose column. 2 The lectin is a glycoprotein with a carbohydrate content of 2.7%. The mol. wt is 392,000 +/- 28,000. Two subunits of identical size (183,000 +/- 3000) are linked by non-covalent bonds. 3. The lectin agglutinates a variety of erythrocytes including human A, B, O red blood cells as well as human lymphocytes. 4. Hemagglutination activity is inhibited best by N,N',N"-triacetylchitotriose, followed by N,N'-diacetylchitobiose, which is half as inhibitory. 5. Lectin activity is constant between pH 5 and 10. Divalent cations are not required for binding reactions. Activity is totally destroyed by heating to 60 degrees C for 30 min. 6. The lectin is precipitated from the extract by 30-40% ammonium sulfate saturation.     

Uridine enhances expression of the fragile X chromosome in human lymphocytes

Summary We have studied the effect of uridine on the expression of fragile X (fra[X]) in lymphocyte cultures established in the folate and thymidine deficient medium TC199. The results indicate that uridine enhances the expression of fra(X) and gives a higher mitotic rate. The excess of uridine during DNA synthesis might further promote the previously suggested cycle of misincorporation and removal of deoxyuridine monophosphate when the pool of deoxythymidine triphosphate is continuously depleted.     

An endogenous ligand for the central sulfonylurea receptor

An endogenous ligand for the rat central sulfonylurea receptor has been evidenced in the rat central nervous system. The characteristics of this ligand (extractibility, non-dialysability, chromatographic behaviour on different media, sensitivity to proteases) indicate that it is a neutral to slightly basic peptide.     

Spring development of a Chlamydomonas population in Lake Nimetön,a small humic forest lake in southern Finland

Biomass development and vertical distribution of a Chlamydomonas population in a small humic forest lake was followed by daily sampling in May-June, 1984. Chlamydomonas dominated the phytoplankton spring bloom, forming 71% of the maximum phytoplankton biomass on 18 May. In early May the outflow rate was high and during the 24 hour period when the maximum rate of surface runoff was recorded (8–9 May), 43% of the Chlamydomonas biomass was flushed out of the lake, which delayed the onset of biomass increase. When surface runoff had slowed down Chlamydomonas biomass started increasing and during wax of the population most cells were < 10 µm in diameter. Population maximum lasted for one day (18 May) and there-after Chlamydomonas biomass decreased towards the end of the study. During wane of the population most cells were > 10 µm in diameter.     

Novel subunit in C4b-binding protein required for protein S binding

C4b-binding protein (C4BP) is a multimeric protein with regulatory functions in the complement system. It also interacts with vitamin K-dependent protein S, which is involved in the regulation of the coagulation system. It has been demonstrated that C4BP consists of seven disulfide-linked, identical 70-kDa subunits, which are arranged to give the molecule a spider-like structure. We now have evidence for the presence of a new subunit in C4BP. On sodium dodecyl sulfate-poly-acrylamide gel electrophoresis it appears as a weakly stainable band with a molecular weight of approximately 45,000. The subunit was isolated by gel filtration in 6 M guanidine hydrochloride of reduced and carboxymethylated C4BP. Its amino-terminal sequence is distinct from previously known protein sequences. The stoichiometry of 45- to 70-kDa subunits was estimated to be 1:9, indicating the presence of one 45-kDa subunit per C4BP molecule. The new subunit was demonstrated to be a disulfide-linked component of the central core of C4BP. It was sensitive to proteolysis by chymotrypsin, and when cleaved the protein S binding ability of C4BP was lost. With protein S bound to C4BP, the 45-kDa subunit was protected from degradation by chymotrypsin, and the protein S binding site remained intact. These data suggest that the new subunit is directly involved in protein S binding.     

Regular and irregular divisions of Lepidoptera spermatocytes as related to the speed of meiotic prophase

Lepidoptera males bear two kinds of meiotic divisions. One is regular (eupyrene) and leads to nucleate, fertilizing spermatozoa. The other (apyrene) shows metaphase I chromosomes clumping together into irregular masses which later split forming daughter cells with unbalanced sets of chromosomes which are eventually extruded from the cells; hence, the spermatids develop into anucleate spermatozoa of unknown function. The apyrene divisions are induced by a haemolymph factor which becomes functional towards pupation. Using incorporation of tritiated thymidine at the premeiotic S-phase as a marker for timing, it was found that the prophase of the apyrene spermatocyte is shorter than that of the eupyrene spermatocyte. It is proposed that meiosis-specific proteins cannot be synthesized during the shortened apyrene prophase and that this is correlated with the irregular chromosome behaviour during the subsequent metaphase-telophase of these spermatocytes.