Stereoselective total synthesis of ceramide Di-, Tri- and tetrahexosides of wheat flour

The first total synthesis of glycosphingolipids isolated from wheat flour has been achieved in a regio- and stereo-controlled manner.Abbreviations THF tetrahydrofuran - DMF dimethylformamide Part 53 in the series Synthetic Studies on Cell Surface Glycans     

Preferential binding of E.coli histone-like protein HU alpha to negatively supercoiled DNA.

Binding specificity of histone-like HU alpha protein to supercoiled DNA was examined by gel retardation assay and chemical probing with OsO4. The latter method was proved to be a unique means for detecting torsional tension restrained in supercoiled plasmid in the presence of HU alpha. It was shown that HU alpha protein has preferential affinity to negatively supercoiled DNA relative to relaxed, nicked and linearized DNAs. There were two modes for binding of HU alpha to the supercoiled DNA: one was the binding associated with topological changes in DNA and the other was relatively strong binding, probably specific to certain particular structures of DNA. It was suggested that HU in vivo interacts preferentially with the regions deformed under torsional stress or with the metabolically active regions along DNA.     

Scavenging of Hydrogen Peroxide in Prokaryotic and Eukaryotic Algae: Acquisition of Ascorbate Peroxidase during the Evolution of Cyanobacteria

Ascorbate (AsA) peroxidase was found in six species of cyanobacteriaamong ten species tested. Upon the addition of H₂¹⁸O₂ to thecells of AsA peroxidase-containing cyanobacteria, ¹⁶O₂ derivedfrom water and ¹⁸O₂ derived from H₂^I8O₂ were evolved in thelight. The evolution of ¹⁶O₂ was inhibited by DCMU and did notoccur in the dark, but ^I8O₂ was evolved even in the dark orin the presence of DCMU. Similar light-dependent evolution of¹⁶O₂ was observed in the cells of AsA peroxidase-containingEuglena and Chlamydomonas. However, the cells of AsA perox-idase-lackingcyanobacteria evolved only ¹⁸O₂ in either the light or dark.Furthermore, the quenching of chlorophyll fluorescence inducedby hydrogen peroxide was observed only in the cells of the AsAperoxidase-containing Synechocystis 6803, and not in the cellsof Anacystis nidulans which lacks AsA peroxidase. Thus, cyanobacteriacan be divided into two groups, those that has and those thatlacks AsA peroxidase. The first group scavenges hydrogen peroxidewith the peroxidase using a photoreductant as the electron donor,and the second group only scavenges hydrogen peroxide with catalase. (Received July 23, 1990; Accepted October 18, 1990)     

Cell-surface hydrophobicity of Candida species as determined by the contact-angle and hydrocarbon-adherence methods

Cell-surface hydrophobicities of six Candida species were studied by two methods: measurement of the contact angle, and partitioning with aqueous-hydrocarbon (n-octane, n-hexadecane and p-xylene) mixtures. C. tropicalis, C. glabrata and C. krusei adhered better to the hydrocarbons than did C. albicans, C. stellatoidea and C. parapsilosis. Contact angles for the less adherent species were smaller than those for the more adherent species. Thus the two methods gave results that were similar overall and indicated that C. tropicalis, C. glabrata and C. krusei have greater cell-surface hydrophobicities than C. albicans, C. stellatoidea and C. parapsilosis.     

N-terminal amino acid sequence of human urinary prokallikrein

The N-terminal amino acid sequences of human urinary prokallikrein and kallikrein have been determined. Their amino acid sequences are as follows. (Formula; see text) The results showed that prokallikrein comprises an additional seven amino acids at the amino terminus of the kallikrein, of which the sequence is (H2N)Ala-Pro-Pro-Ile-Gln-Ser-Arg(COOH). Comparison of the structure of this peptide with those of other proteins revealed extensive sequence identity with the propeptide portions of rat and mouse tissue kallikreins, that were predicted from the preproenzyme-encoded nucleotide sequences. The amino acid sequence of the peptide was also highly homologous to that of the propeptide portion of EGF-binding protein, that was predicted from the nucleotide sequence, and that of the alpha-subunit of NGF. The N-terminal amino acid sequence of kallikrein was completely identical to the reported one (Lottspeich, F., et al. (1979) Hoppe-Seyler's Z. Physiol. Chem. 360, 1947-1950) and shows considerable amino acid sequence homology with the porcine and rat pancreatic kallikreins. As far as the present results are concerned, it is strongly indicated that the inactive kallikrein in human urine is a tissue type prokallikrein which is activated on the release of the N-terminal peptide consisting of seven amino acids.     

A spin-label study on human high density lipoprotein

Human plasma high density lipoproteins (HDL) have been labeled with N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)maleimide (NEM-TEMPO). The spin-labeled HDL exhibited an ESR spectrum containing signals of both strongly immobilized and weakly immobilized components by the reaction with a high concentration of NEM-TEMPO, while an ESR spectrum containing only signals of a strongly immobilized component range between 4 degrees C and 37 degrees C, the signal height of the strongly immobilized component exhibited reversible temperature-dependent changes, whereas that of the weakly immobilized component changed irreversibly at temperatures above 25 degrees C. The activation energy of the irreversible change was estimated to be 26 kcal per mol. The strongly immobilized component was derived from NEM-TEMPO which modified apolipoprotein A-I covalently, while the weakly immobilized component was derived from NEM-TEMPO noncovalently bound to HDL. The rate of binding of NEM-TEMPO to either the strongly binding or weakly binding sites and the number of the strongly binding sites in apolipoprotein A-I were estimated to be 125 M-1.day-1 and 1.78, respectively. The binding of NEM-TEMPO to the strongly binding sites was suppressed greatly by pretreatment of HDL with 2,4,6-trinitrobenzene sulfonic acid (TNBS). The slow reaction and suppression with TNBS suggest that NEM-TEMPO binds to some amino acid residue, probably a lysine residue, in apoprotein A-I. The strongly immobilized and weakly immobilized components were reduced almost completely by ascorbate at the same rate, 0.048 min-1 at pH 7.4 and at 4 degrees C.     

Radiation-induced peroxidation of lipid dissolved in organic solvent and its inhibition by alpha-tocopherol and cepharanthine

Effects of cepharanthine and alpha-tocopherol on radiation-induced peroxidation of lipids dissolved in methanol(MeOH)-chloroform (CHCl3)-H2O(v/v, 2/1/0.8) were examined. alpha-Tocopherol strongly inhibited radiation-induced peroxidation of lipids dissolved in MeOH-CHCl3-H2O. However, cepharanthine exhibited a weak inhibitory action in this system. The change in the absorption spectrum of alpha-tocopherol and cepharanthine by X-irradiation was measured. The reagents were dissolved in 95% EtOH acidified with 20 mM HCl and in MeOH-CHCl3-H2O. alpha-Tocopherol exhibited the change in its absorption spectrum in both systems, and seemed to be oxidized at a high rate by free radicals. However, cepharanthine slightly exhibited the change in its spectrum in MeOH-CHCl3-H2O, but not in acidified EtOH.     

Cell lines from imaginal discs ofDrosophila melanogaster

Summary New cell lines, designated as ML-DmDl≈10, were established from dissociated imaginal discs ofDrosophila melanogaster. The culture medium was prepared by mixing in a 1:1 ratio Cross and Sang’s M3(BF) medium, supplemented with 10% heat inactivated fetal bovine serum (FBS), with the supernatant of a primary embryonic cell culture made in the M3(BF) medium and supplementing this mixture with insulin. One cell line was established in the medium containing larval hemolymph instead of the primary culture supernatan, and another was established in fresh M3(BF) medium supplemented with insulin and FBS. In these mediums, imaginal disc cells first formed aggregates and cellular vesicles within a few weeks followed by the proliferation of thin-layered cells around them after about 1 mo. Ten cell lines have so far been established from two kinds of imaginal discs and disc mixtures. The ploidy of these cell lines was predominantly diploid. Population doubling time was about 50 to 70 h at 3 to 10 mo. after initiation of the culture. When the cell aggregates formed in vitro were implanted in metamorphosing larvae, they differentiated at high frequency into adult cuticular strutures in the early phase of the primary culture. This differentiation of aggregates was also observed, though at low frequency, in a culture maintained by dilution-transfer for 6 to 15 mo. in vitro.