Non-cognizable ribonucleotide, 2′AMP, binds to a mutant ribonuclease T1 (Y45W) at a new base-binding site but not at the guanine-recognition site

Complex of a mutant ribonuclease T₁ (Y4SW) with a non-cognizable ribonucleotide, 2′AMP, has been determined and refined by X-ray diffraction at 1.7 Å resolution. The 2′AMP molecule locates at a new base-binding site which is remote from the guanine-recognition site, where 2′GMP was found to be bound. The nucleotide adopts the anti conformation of the glycosidic bond and C3′-exo sugar pucker. There exists a single hydrogen bond between the adenine base and the enzyme, and, therefore, the site found is apparently a non-specific binding site. The results indicate that the binding of 2′AMP to the guanine-recognition site is weaker than that to the new binding site.     

Infrequent but significant p21/WAF1/CIP1 gene alteration in synchronous or metachronous transitional cell carcinoma

p53 inducible cyclin dependent kinase inhibitor, p21/WAF1/CIP1(p21), played a pivotal role for G1 arrest when cells received genotoxic stimuli. p21 could be a putative tumor suppressor gene, since its dysfunction may lead to accumulation of genomic alteration. We investigated the p21 and p53 status using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and immunohistochemical analyses, in eight patients who had synchronous or metachronous urothelial tumors. Loss of heterozygosity (LOH) of p21 gene was detected in one coincidental tumor in one case. p21 positive cells were detected by immuno-histochemical staining in all tumors in one case, and in one coincidental tumor in two cases. Among p21 positive cells in these three cases, no p53 mutations were detected, whereas no p21 positive cells were detected in other cases with a p53 mutation. These findings suggested that in transitional cell carcinoma (TCC) p21 gene mutation is infrequent like the p53 gene mutation, but that LOH might be important in the inactivation of p21.     

Phylogenetic Analysis of the Genus Bifidobacterium and Related Genera Based on 16S rDNA Sequences

The 16S rRNA gene sequences were determined for type strains of 21 Bifidobacterium species. A phylogenetic tree was constructed using the determined sequences and sequences from DNA databases, which contain the sequences of 11 type strains of Bifidobacterium species and 11 strains of related genera. All species of the genus Bifidobacterium and Gardnerella vaginalis ATCC 14018 belonged to a cluster phylogenetically distinct from the other genera. The cluster was divided into two subclusters: subcluster 1 composed of most species of Bifidobacterium and G. vaginalis, and subcluster 2 consisting of two species, B. denticolens and B. inopinatum; both of which were isolated from human dental caries. In the genus Bifidobacterium, four groups of species are known to be moderately to highly related by DNA-DNA hybridization. The four groups of species exhibited more than 99% similarity among their 16S rDNA sequences within each group. These results indicated that species with around 99% or more similarity in their 16S rDNA sequences should be confirmed for species identities.     

Allo-cedrol: A new tricarbocyclic sesquiterpene alcohol

The structure of a new tricarbocyclic sesquiterpene alcohol, named allo-cedrol, was determined by relation with β-acoradiene. The enantiomer of prezizaene and several compounds were obtained from the p-bromobenzenesulfonate of allo-cerol.     

Counterflow efflux of thiamin in Escherichia coli

A resting cell of Escherichia coli lacking thiamin kinase incorporated external thiamin with an energy-dependent counterflow efflux (C-efflux). This C-efflux could be separated from an energy-dependent exit by a selective inhibition of exit by $\text{2 · 10}{}^{\mathrm{?2}}\text{M}$ NaN₃. The extracellular thiamin could be replaced by thiamin diphosphate, resulting in the same rate of C-efflux, but the rate of C-efflux of intracellular thiamin diphosphate against the external thiamin was markedly low. This low rate of C-efflux of thiamin diphosphate could explain the higher accumulation of the compound than that of free thiamin in the thiamin-kinase-defective mutant as well as in its wild-type parent. Basic characteristics of free thiamin uptake and exit in E. coli W mutant were compared with those reported in K 12 mutant: a marked difference existed in the rate of exit. The low rate of exit in E. coli W 70-23-102 was inferred as the reason for the absence of an overshoot phenomenon of thiamin uptake in this strain.     

The molecular structure of cyclonucleotide hexamer, CoGoCoGoCoGo having a high-anti conformation

Cyclonucleotide hexamer, CoGoCoGoCoGo, was synthesized and crystallized as orthorhombic with space group C222(1), and unit cell dimensions: a = 48.30, b = 41.53, and c = 31.76A. The X-ray diffraction data up to 1.8A resolution were collected, and the crystal structure analysis by molecular replacement technique is now in progress using two energetically adequate left-handed helical models, which are obtained by conformational energy calculation.     

Light and electron microscopic studies in the cerebellum of "trembler" mutant of chickens

Light and electron microscopic analyses of the cerebellar cortex were carried out in inbred trembler mutants of Barred Plymouth Rock (BPR), crossbred trembler (CBT) and crossbred normal (CBN) chickens 6 and 34 days after hatching. The size of the cerebellum of BPR was markedly reduced, but the pattern of fissures appeared essentially normal. Both the molecular and granular layer in the BPR were much thinner than in the CBT and CBN. Ectopic Purkinje cells were observed in the molecular and granular layers, and even in the medulla. The Purkinje cells were more intensely stained in Nissl preparations than those of CBT and CBN. By 34 days, most of the Purkinje cells, which were reduced in size, contained spherical and swollen mitochondria, an undeveloped Golgi apparatus, and many stacks of rough endoplasmic reticulum (ER). The basket cells were reduced in size and exhibited a decrease in stacks of rough ER and an increase in the area of the Golgi apparatus. They contained the swollen mitochondria that were observed in the Purkinje cells. The granule cells were reduced in size, and showed a decrease in the number of cell organelles. The disorder appeared to be intrinsic to Purkinje cells since many types of degeneration were observed in the BPR Purkinje cells.     

Structure and biosynthesis of prototokoronin in tissue cultures of Dioscorea tokoro

The furostanol glucoside prototokoronin (1) has been isolated from tissue cultures of Dioscorea tokoro M., and labelled yonogenin and diosgenin were incorporated into (1). Sapogenin and protosaponin are thus seen to be interconvertible in tissue cultures.     

Introns and reading frames: correlation between splicing sites and their codon positions

Computer analyses of the entire GenBank database were conducted to examine correlation between splicing sites and codon positions in reading frames. Intron insertion patterns (i.e., splicing site locations with respect to codon positions) have been analyzed for all of the 74 codons of all the eukaryote taxonomic groups: primates, rodents mammals, vertebrates, invertebrates, and plants. We found that reading frames are interrupted by an intron at a codon boundary (as opposed to the middle of a codon) significantly more often than expected. This observation is consistent with the exon shuffling hypothesis, because exons that end at codon boundaries can be concatenated without causing a frame shift and thus are evolutionarily advantageous. On the other hand, when introns interrupt at the middles of codons, they exist in between the first and second bases much more frequently than between the second and third bases, despite the fact that boundaries between the first and second bases of codons are generally far more important than those between the second and third bases. The reason for this is not clear and yet to be explained. We also show that the length of an exon is a multiple of 3 more frequently than expected. Furthermore, the total length of two consecutive exons is also more frequently a multiple of 3. All the observations above are consistent with results recently published by Long, Rosenberg, and Gilbert (1995).