Cloning and over-expression of thermostable Bacillus sp. YM55-1 aspartase and site-directed mutagenesis for probing a catalytic residue.

A thermostable aspartase gene (aspB) from Bacillus sp. YM55-1 was cloned and the gene sequenced. The aspB gene (1407 bp ORF) encodes a protein with a molecular mass of 51 627 Da, consisting of 468 amino-acid residues. An amino-acid sequence comparison revealed that Bacillus YM55-1 aspartase shared 71% homology with Bacillus subtilis aspartase and 49% with Escherichia coli and Pseudomonas fluorescens aspartases. The E. coli TK237/pUCASPB strain, which was obtained by transforming E. coli TK237 (aspartase-null strain) with a vector plasmid (pUCASPB) containing the cloned aspB gene, produced a large amount of the enzyme corresponding to > 10% of the total soluble protein. The over-expressed recombinant enzyme (native molecular mass: 200 kDa) was purified effectively and rapidly using heat treatment and affinity chromatography. In order to probe the catalytic residues of this enzyme, two conserved amino-acid residues, Lys183 and His134, were individually mutated to alanine. Although the tertiary structure of each mutant was estimated to be the same as that of wild-type aspartase in CD and fluorescence measurements, the Lys183Ala mutant lost its activity completely, whereas His134Ala retained full activity. This finding suggests that Lys183 may be involved in the catalytic activity of this thermostable Bacillus YM55-1 aspartase.

[A clinico-pathological study of 69 cases after total cystectomy].

A clinico-pathological study was conducted on 69 patients with bladder cancer who underwent total cystectomy. The one, three and five-year actuarial survival rates for the 69 patients were 73.3%, 48.6% and 44.1%, respectively. Survival rate was not significantly associated with sex, the number of tumors or the size of tumors. The survival rate in those aged 70 years or more was slightly worse than in those who were much younger. Patients with papillary tumors had a more favorable survival rate than those with non-papillary tumors but we could find no significant difference between those with pedunculated tumors and those with sessile tumors. The actuarial 5-year survival rates by grade were 71.9% in G1 + G2, 22.6% in G3 and 32% in non-transitional cell carcinoma; the rates by stage were 86.5% in pTa + pT1, 85.7% in pT2, 20.8% in pT3a, 18.2% in pT3b and 0% in pT4. When the stage reached pT3a, the survival rate fell remarkably. The rate of INF alpha (93.8%) was significantly better than that of INF beta (28.1%) and INF gamma (15.2%). The rate of ly0 (76.2%) was also significantly better than that of ly1 (25.5%) and ly2 (18.8%). There was no significant difference in survival between v(-) (50.7%) and v(+) (25.9%). We could find no significant difference between patients who underwent pelvic lymph nodes dissection and those who did not.(ABSTRACT TRUNCATED AT 250 WORDS)

[The application of multivariate analysis to the radioallergosorbent test (RAST)].

Multivariate statistical analyses using principal component analyses (PCA) and Mahalanobis' method were applied to the quality control of radioallergosorbent test (RAST). Mites and foods allergen were analyzed by PCA method, and foods allergen were analyzed by Mahalanobis' method. The vast differences in the distribution on titers among the mites and the foods were observed. To evaluate the results of a test and the quality of kits for test statistically, it should be better to use multivariate analyses such as PCA or Mahalanobis' method.

Paratesticular liposarcoma: coexistence with urinary bladder cancer.

A 58-year-old man with liposarcoma originating from the right paratesticular area was reported. Histologically, liposarcoma showed a well-differentiated sclerosing type. Coexistence of transitional cell carcinoma of the urinary bladder and liposarcoma was also revealed. The cases of paratesticular liposarcoma in the literature are briefly reviewed.

[Bladder carcinoma with sarcomatous changes].

A 78-year-old man underwent surgery to remove a bladder tumor. The bladder contained two solid pedunculated tumors, which existed independently. Histopathologic examination revealed that of most the tumor was composed of spindle-shaped sarcomatous tissue sometimes containing multinucleated giant cells. Transitional cell carcinoma (Grade 3) was also found at the stalk of the tumor with squamous metaplasia. The squamous metaplasia, which gradually changed into sarcomatous tissue, was also found at many parts of the tumor, but no definite transition of the transitional cell carcinoma into sarcomatous tissue was noted. Therefore, the sarcomatous change of carcinoma in our case was brought about by way of squamous metaplasia of transitional cell carcinoma. The tumor could be given the diagnosis of so-called "carcinosarcoma". The pathologic significance of "carcinosarcoma" was briefly discussed.