The specificity of chitinase C-1 of Streptomyces griseus HUT 6037 for the hydrolysis of the ␤-1,4-glycosidic linkages in partially acetylated chitosan is different from that of other microbial chitinases. In order to study the primary structure of this unique chitinase, the chiC gene specifying chitinase C-1 was cloned and its nucleotide sequence was determined. The gene encodes a polypeptide of 294 amino acids with a calculated size of 31.4 kDa. Comparison of the amino acid sequence of the deduced polypeptide with that of other proteins revealed a C-terminal catalytic domain displaying considerable sequence similarity to the catalytic domain of plant class I, II, and IV chitinases which form glycosyl hydrolase family 19. The N-terminal domain of the deduced polypeptide exhibits sequence similarity to substrate-binding domains of several microbial chitinases and cellulases but not to the chitin-binding domains of plant chitinases. The previously purified chitinase C-1 from S. griseus is suggested to be generated by proteolytic removal of the N-terminal chitin-binding domain and corresponds to the catalytic domain of the chitinase encoded by the chiC gene. High-performance liquid chromatography analysis of the hydrolysis products from N-acetyl chitotetraose revealed that chitinase C-1 catalyzes hydrolysis of the glycosidic bond with inversion of the anomeric configuration, in agreement with the previously reported inverting mechanism of plant class I chitinases. This is the first report of a family 19 chitinase found in an organism other than higher plants.Chitinases (EC 3.2.1.14) are glycosyl hydrolases which catalyze the degradation of chitin, an insoluble linear ␤-1,4-linked polymer of N-acetylglucosamine. Recently, chitinases have been receiving renewed attention because of their possible application for the biological control of chitin-containing organisms and also for the exploitation of natural chitinous materials. They are present in a wide range of organisms including bacteria, insects, viruses, plants, and animals and play important physiological and ecological roles. Chitinases so far sequenced are classified in two different families (namely, families 18 and 19) of the classification of glycosyl hydrolases based on amino acid sequence similarities (13,14). Family 18 contains chitinases from bacteria, fungi, viruses, and animals and class III and V chitinases from plants. On the other hand, class I, II, and IV plant chitinases belong to family 19, and this family solely comprises chitinases of plant origin. The two different families of chitinases display no sequence similarities with each other and have different three-dimensional structures (5).Oligosaccharides produced from partially acetylated chitosan by microbial chitinases have been previously studied in an attempt to clarify their specificity for ␤-1,4-N-acetylglucosaminic versus ␤-1,4-glucosaminic linkages (26, 27, 30). Except for chitinase C-1 from Streptomyces griseus HUT 6037, all of the examined microbial chitinases hydrolyzed GlcNAc-GlcNA...

Our results suggest that APP duplication should be considered in patients with EO-FAD in various ethnic groups, and that increased APP mRNA expression level owing to APP duplication contributes to AD development.

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Background. Gonadotropin‐releasing hormone (GnRH) analogs have been used in the therapy of the endocrine‐dependent cancers. The authors attempted to determine the frequency with which Gn‐RH receptor (Gn‐RHR) is present in gynecological cancers. Methods. Experiments were performed on gynecologic tumors that had been surgically removed and their cloned cell lines. Gn‐RHR was characterized by [3H]Gn‐RH binding to plasma membrane preparations. Gn‐RHR messenger ribonucleic acid was determined by reverse transcription‐polymerase chain reaction using oligonucleotide primers synthesized according to the published human Gn‐RHR sequence. Results. High affinity binding sites with nanomolar range of Kd and Gn‐RHR mRNA were detected in a high proportion (over 90%) of the specimens from endometrium (6 of 6) and endometrial carcinomas (16 of 17), myometrium (6 of 6) and myomas (4 of 5), epithelial carcinoma (21 of 23), and stromal tumors (3 of 3) of the ovary. There was no substantial Gn‐RHR in cervical carcinomas or germ cell‐derived tumors of the ovary. Cloned cell lines gave identical results to those obtained in their respective mother tumors. Conclusions. We detected Gn‐RHR in a wide range of the carcinomas and tissues originating from the endometrium and ovary, but not in the uterine cervix or germ cell‐derived tumors. The expression of Gn‐RH receptor raises the possibility that Gn‐RH may play a direct regulatory role in the growth of these carcinomas, and provides a possible point of attack for therapeutic approaches using Gn‐RH analogs in these malignancies.

Background/Aims: Recent clinical applications suggest a beneficial effect of gonadotropin-releasing hormone analog (GnRHa) as a gonadal protector from chemotherapy-induced premature ovarian failure. This study aimed to determine cellular mechanisms involved in the protective action of GnRHa against granulosa cell damage caused by doxorubicin. Methods: Granulosa cells were obtained by ultrasound-guided follicular aspiration from patients undergoing in vitro fertilization, and screened for GnRH receptor expression prior to analyses. The cellular function was assessed by measuring the conversion of exogenously supplied androstenedione to estradiol-β (E2) in response to follicle-stimulating hormone (FSH) (1 µM). Results: Exposing to doxorubicin for 12 h before FSH stimulation caused a concentration-dependent inhibition of the E2 secretion to a minimum level of 20% of control. When the cells were incubated with a GnRHa for 12 h before and during exposure to doxorubicin, granulosa cells produced an equal level of E2 to that of control cells. The protective action of GnRHa was dose-dependent; a half-maximal effect occurred at 10 nM. Preincubation with GnRHa alone had no effect on FSH-induced E2 production. Conclusion: These findings demonstrate that a GnRHa may retard doxorubicin-induced granulosa cell damage, suggesting an additional GnRH activity to protect the gonads during chemotherapy through GnRH receptor-mediated mechanism(s).