In the course of our screening of /?-glucosidase inhibitor, a culture filtrate of a mushroom, Phellinus sp. strongly inhibited the enzymeactivity. The active substance wasisolated through charcoal separation, column chromatography and crystallization. Spectroscopic and crystallographic analysis revealed that it had a novel cyclitol structure, (15f,2i?,35f,4/?,57?,6i?)-5-hydroxymethyl-7oxabicyclo[4, l ,0]heptane-2,3,4-triol, and we named it cyclophellitol. It inhibited almond-derived /?-glucosidase with an IC50 of 0.8 ug/ml. 49 /?-Glucosidase inhibitors such as castanospermine and 1-deoxynojirimycin have been reported to inhibit syncytium formation and infection of human immunodeficiency virus (HIV), possibly by perturbing gpl20-linked glycan structurelf2). Castanospermine is also known to suppress experimental metastasis possibly by changing the saccharide structure on tumour cell surface3*. Therefore, glucosidase inhibitors may inhibit HIV infection and metastasis. Consequently we screened culture filtrates of microorganisms for inhibitory activity against^-glucosidase. Amonga thousand strains of bacteria, Actinomycetes and mushrooms,we found that a culture filtrate of a mushroomstrain, Phellinus sp., showedinhibitory activity against almond^-glucosidase. Isolation and structure determination of the active principle, established that it was a novel compound. Wehave namedit cyclophellitol. Materials and Methods General /7-Nitrophenyl-^-D-glucopyranoside and almond /?-glucosidase were purchased from Sigma. NMR spectra were recorded on a Jeol JNM-GX400. The MSspectra were taken by a Hitachi M-80H spectrometer. The mp was measured by the micro mp apparatus, MP-S3(Yanagimoto). The UVand IR spectra were measured by a Hitachi 220S and a 260-10 spectrophotometer, respectively. Optical rotations were taken by a Perkin-Elmer 241 polarimeter using micro-cell (light path 10 cm). #-Glucosidase Assay The enzyme activity was assayed by the method described by Saul et al.4\ with slight modifications.
VIP36, an intracellular lectin that recognizes high mannose-type glycans (Hara-Kuge, S., Ohkura, T., Seko, A., and Yamashita, K. (1999) Glycobiology 9, 833-839), was shown to localize not only to the early secretory pathway but also to the plasma membrane of MadinDarby canine kidney (MDCK) cells. In the plasma membrane, VIP36 exhibited an apical-predominant distribution, the apical/basolateral ratio being ϳ2. Like VIP36, plasma membrane glycoproteins recognized by VIP36 were found in the apical and basolateral membranes in the ratio of ϳ2 to 1. In addition, secretory glycoproteins recognized by VIP36 were secreted ϳ2-fold more efficiently from the apical membrane than from the basolateral membrane. Thus, the apical/basolateral ratio of the transport of VIP36-recognized glycoproteins was correlated with that of VIP36 in MDCK cells. Upon overproduction of VIP36 in MDCK cells, the apical/basolateral ratios of both VIP36 and VIP36-recognized glycoproteins were changed from ϳ2 to ϳ4, and the secretion of VIP36-recognized glycoproteins was greatly stimulated. In contrast to the overproduction of VIP36, that of a mutant version of VIP36, which has no lectin activity, was of no effect on the distribution of glycoproteins to apical and basolateral membranes and inhibited the secretion of VIP36-recognized glycoproteins. Furthermore, the overproduction of VIP36 greatly stimulated the secretion of a major apical secretory glycoprotein of MDCK cells, clusterin, which was found to carry at least one high mannose-type glycan and to be recognized by VIP36. In contrast to the secretion of clusterin, that of a non-glycosylated apical-secretion protein, galectin-3, was not stimulated through the overproduction of VIP36. These results indicated that VIP36 was involved in the transport and sorting of glycoproteins carrying high mannose-type glycan(s).Newly synthesized secretory and membrane proteins exit from the ER 1 in transport vesicles targeted to the Golgi apparatus. Vesicular transport through the Golgi is often accompanied by post-translational modifications, such as glycosylation of cargo proteins until they have reached the trans-Golgi Network. In the trans-Golgi Network, proteins are sorted into vesicles bound for different destinations including the plasma membrane, the endosome/lysosome, and secretory granules. The protein sorting has been one of the most interesting issues in the study of vesicular protein traffic processes, but its molecular mechanisms remain largely unresolved.It has been recently demonstrated that intracellular lectins play important roles in vesicular transport: for example, mannose-6-phosphate receptor (1) as a receptor recognizing the marker for lysosomal enzymes, calnexin (2, 3) and calreticulin (4) as molecular chaperones, and ERGIC-53 (5) possibly as a transport cargo receptor. ERGIC-53 is an intermediate compartment marker (6), and it is identical to MR60, a mannosespecific membrane lectin (7) with a carbohydrate-binding domain homologous to that of lectins of leguminous plants (8). The N-term...
This paper describes a novel mutant mouse that has been spontaneously derived from the Snell's dwarf (DW/J) mouse. It was named the 'growth-retarded mouse' because of a characteristic growth pause followed by the delayed onset of pubertal growth. The onset of the increase in pituitary GH content that normally occurs concomitant with pubertal growth was also delayed in the growth-retarded mice. The serum concentration of thyroxine was very low in these mice from the neonatal period through adulthood, and a supplement of tri-iodothyronine was effective in shortening the growth pause and commencing the suppressed pubertal growth. Histological and immunohistochemical studies revealed that the anterior pituitary gland of the growth-retarded mouse contains clustered unusual chromophobic cells which are not reactive to various antisera against anterior pituitary hormones and the gland becomes enlarged with age. Breeding data indicated that these characteristics of the mice show an autosomal recessive inheritance and the gene responsible was designated as 'grm'. Partial linkage analysis utilizing microsatellite polymorphism demonstrated that the grm gene does not identify with the lit or hyt genes. Based on comparison of the hormonal status and growth pattern between growth-retarded, dwarf and normal mice, we have suggested the existence of a mutual interaction, possibly positive feedback regulation, between the pituitary and thyroid glands, that develops or matures the hormonal network which is responsible for rapid somatic growth and metabolic changes at puberty in mice.
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