Crystal structures and inhibitor binding properties of plant class V chitinases: the cycad enzyme exhibits unique structural and functional features.

A class V (glycoside hydrolase family 18) chitinase from the cycad Cycas revoluta (CrChiA) is a plant chitinase that has been reported to possess efficient transglycosylation (TG) activity. We solved the crystal structure of CrChiA, and compared it with those of class V chitinases from Nicotiana tabacum (NtChiV) and Arabidopsis thaliana (AtChiC), which do not efficiently catalyze the TG reaction. All three chitinases had a similar (α/ß)8 barrel fold with an (α + ß) insertion domain. In the acceptor binding site (+1, +2 and +3) of CrChiA, the Trp168 side chain was found to stack face-to-face with the +3 sugar. However, this interaction was not found in the identical regions of NtChiV and AtChiC. In the DxDxE motif, which is essential for catalysis, the carboxyl group of the middle Asp (Asp117) was always oriented toward the catalytic acid Glu119 in CrChiA, whereas the corresponding Asp in NtChiV and AtChiC was oriented toward the first Asp. These structural features of CrChiA appear to be responsible for the efficient TG activity. When binding of the inhibitor allosamidin was evaluated using isothermal titration calorimetry, the changes in binding free energy of the three chitinases were found to be similar to each other, i.e. between -9.5 and -9.8 kcal mol(-1) . However, solvation and conformational entropy changes in CrChiA were markedly different from those in NtChiV and AtChiC, but similar to those of chitinase A from Serratia marcescens (SmChiA), which also exhibits significant TG activity. These results provide insight into the molecular mechanism underlying the TG reaction and the molecular evolution from bacterial chitinases to plant class V chitinases.

A CASE REPORT OF PANCREATIC METASTASIS FROM PROSTATE CANCER.

We report a case of pancreatic metastasis from prostate cancer. A 65 year-old man developed gross hematuria, and was found to have a markedly elevated PSA and abnormal CT findings indicating mass lesions in bilateral lungs, and pancreatic head, and osteoblastic lesions in pelvic bone. He was referred to the department of Urology and Gastroenterology at Seirei Hamamatsu General Hospital. Digital rectal examination revealed a hen's egg-sized, stony-hard prostate. A systematic needle biopsy of the prostate demonstrated adenocarcinoma (Gleason score 4+4=8). We diagnosed multiple lung and bone metastases from prostate cancer, and suspected that a primary pancreatic ductal carcinoma. The patient was started on an androgen deprivation therapy (ADT) with bicalutamide and degarelix. After 4 months from the initiation of ADT, an MRI of the pancreas showed decreased size of the pancreatic head mass from 18 mm to 7 mm in diameter. We concluded that the pancreatic head mass was metastasis from prostate cancer, and currently continue ADT. Pancreatic metastasis from prostate cancer is very rare, and further accumulation of cases will be required.

Mutation strategies for obtaining chitooligosaccharides with longer chains by transglycosylation reaction of family GH18 chitinase.

Enhancing the transglycosylation (TG) activity of glycoside hydrolases does not always result in the production of oligosaccharides with longer chains, because the TG products are often decomposed into shorter oligosaccharides. Here, we investigated the mutation strategies for obtaining chitooligosaccharides with longer chains by means of TG reaction catalyzed by family GH18 chitinase A from Vibrio harveyi (VhChiA). HPLC analysis of the TG products from incubation of chitooligosaccharide substrates, GlcNAc(n), with several mutant VhChiAs suggested that mutant W570G (mutation of Trp570 to Gly) and mutant D392N (mutation of Asp392 to Asn) significantly enhanced TG activity, but the TG products were immediately hydrolyzed into shorter GlcNAc(n). On the other hand, the TG products obtained from mutants D313A and D313N (mutations of Asp313 to Ala and Asn, respectively) were not further hydrolyzed, leading to the accumulation of oligosaccharides with longer chains. The data obtained from the mutant VhChiAs suggested that mutations of Asp313, the middle aspartic acid residue of the DxDxE catalytic motif, to Ala and Asn are most effective for obtaining chitooligosaccharides with longer chains.

[Laparoscopic resection of a retroperitoneal müllerian cyst: a case report].

A 51-year-old woman had a cystic mass in the retroperitoneal space, below the left kidney, which was incidentally detected at a medical check-up. The size of the mass was 6 cm in diameter, which was similar to that obtained by magnetic resonance imaging 4 years ago. We followed the case and found that the mass was slightly enlarged a year later. Because malignancy could not be ruled out, we performed a laparoscopic tumor excision. Histologically, the cyst was diagnosed as a Müllerian cyst, and there was no evidence of malignancy. Retroperitoneal Müllerian cyst is a rare tumor. Sixteen cases have been reported previously and this is the fourth case of a laparoscopic excision.

A case of prostatic metastasis from non-seminomatous testicular cancer.

Introduction: Prostatic metastasis from testicular cancer is extremely rare, with only 10 reported cases, all of which were diagnosed as relapse. Herein, we report the case of a patient with concurrent testicular cancer and prostatic metastasis. Case presentation: A 57-year-old man presented at our emergency department with urinary retention. A painless mass was found in the right scrotum, and computed tomography showed lung, mediastinal, and liver metastases, and an enlarged prostate. Tumor markers were measured in 2057 U/L lactate dehydrogenase, 2460 mIU/mL human chorionic gonadotrophin, 1303 ng/mL alpha-fetoprotein, and 1.51 ng/mL prostate specific antigen. An orchiectomy and biopsy were performed; the pathological results showed immature teratomas, embryonal carcinomas, choriocarcinomas, and seminomas in the testis, and embryonal carcinomas in the prostate, liver, and mediastinum. The patient refused chemotherapy and died 3 months following diagnosis. Conclusion: Prostatic metastasis should be considered in cases of dysuria or prostate enlargement in testicular cancers.

[LONG TERM ADMINISTRATION OF RELATIVE DOSE INTENSITY-ADJUSTED CABAZITAXEL FOR METASTATIC CASTRATION-RESISTANT PROSTATE CANCER: A CASE REPORT].

We report the case of a 74-year-old man with metastatic castration-resistant prostatic cancer (CRPC), who underwent treatment with cabazitaxel. Initially, he underwent docetaxel treatment for 2 years and exhibited severe neuropathy in his hands caused by its toxicity. As a result, we replaced docetaxel with cabazitaxel. On receiving 100% of the dose of cabazitaxel in the first course, febrile neutropenia (FN) was observed on the seventh day. However, he soon recovered from the FN and we began relative dose intensity (RDI) treatment with an adequate dose-volume and interval of treatments. He was successfully administered 51 cabazitaxel treatment courses without severe adverse effects. Cabazitaxel is a highly effective drug used as second-line chemotherapy following docetaxel, and it causes fewer adverse effects compared with docetaxel. Cabazitaxel may be a suitable alternative for outpatient treatment. Given that the patient in this case had a long overall survival of more than 3.5 years and received over 50 courses of cabazitaxel, it is crucial for RDI to be taken very seriously.

alpha-Casein improves the gel properties of dried egg white.

The effects of addition of alpha-casein (alpha-CN) to dried egg white (DEW) were investigated by measuring transparency, hardness, and water-holding capacity (WHC) of the heat-induced gels. A DEW concentration of 8% (w/w) was required for formation of a self-supporting gel following heating at 80 degrees C for 20 min at pH 7. Solutions of alpha-CN, even up to a protein concentration of 12% (w/w), did not gel under the same conditions. The addition of alpha-CN (0.5-4%) to 8% DEW caused the increase in gel hardness gels, as compared with DEW gels alone at a total amount of protein concentrations, and the mixed gels became transparent with the increase of added alpha-CN concentrations. The 10% mixed protein solutions of alpha-CN (3-6%) and DEW (4-7%) formed transparent gels, although each protein did not gel individually at their protein concentrations. Mixture with 2:8 mixing ratio of alpha-CN to DEW at a total protein concentration of 10% showed synergistic effects in improving DEW gel properties above pH 7 and below 25 mM NaCl. The improvements (hardness, transparency, and WHC) of DEW gel by alpha-CN seem to be caused mainly by the inhibition of alpha-CN against heat coagulation of DEW protein.

Ability of alphas-Casein to suppress the heat aggregation of ovotransferrin.

The effects of alphas-casein on heat aggregation of ovotransferrin (OT) were studied by heating at 80 degrees C for 20 min in 10 mM phosphate buffer, pH 7.0. The heat interactions between alphas-casein and OT were followed by turbidity development and polyacrylamide gel electrophoresis. We found that alphas-casein can effectively suppress the heat-induced aggregation of heat-labile OT. The suppressive ability of alphas-casein was reduced by the presence of NaCl on heating. Dephosphorylated alphas-casein had less ability to suppress the aggregation of OT than native alphas-casein. Our results indicate that alphas-casein interacts with the heat-denatured OT through its exposed hydrophobic surface and phosphoserine residue. Such interactions seem to be important in helping to suppress the aggregation of heated OT. The suppressive effects of alphas-casein on heat aggregation of OT would be partially ascribed to the formation of transparent gel from egg white by the addition of alphas-casein.