Taurocholic acid does not induce apoptosis in HCT116 cells regardless of its intracellular concentration

Hydrophobic bile acids but not hydrophilic bile acids induce apoptosis in HCT116 cells. We expressed sodium-dependent bile acid transporters in HCT116 cells, and the intracellular concentration of hydrophilic bile acids increased to that of the hydrophobic bile acids. But no sign of apoptosis was observed, which suggests a hydrophobic-bile acid-specific mechanism for the induction of apoptosis in HCT116 cells.     

Kinetics of selective acylation of sn-glycerol 3-phosphate in the synthesis of phospholipid molecular species.

The kinetics of the sn-glycerol 3-phosphate acyltransferase [EC 2.3.1.15] reaction support the view that the selective acylation primarily depends on the differences in the affinity of the enzyme for acyl-CoAs.     

Three-dimensional orientations of talar articular surfaces in humans and great apes

The morphology of the talus prescribes relative positions and movements of the calcaneus and navicular with respect to the tibia, hence determining the overall geometry, mobility and function of the foot that mechanically interacts with environments. Clarifying the variations of the articular surface orientations of the talus in humans and extant great apes is therefore of importance in understanding the evolution of bipedal locomotion in the human lineage. The aim of this study is to clarify the three-dimensional orientations of three articular surfaces of the talus (superior, posterior calcaneal and navicular articular surfaces) by means of the newly proposed surface approximation method. Thirty-two tali in humans, chimpanzees, gorillas and orangutans were scanned using a three-dimensional noncontact digitizer, and the articular surfaces were then approximated using a paraboloid or a plane to calculate the orientations of the surfaces with respect to the body of the talus. The results quantitatively demonstrated that the superior articular surfaces in humans were relatively more parallel with the horizontal plane of the talar body, while those in apes were more medially oriented. Furthermore, the cylindrical axis defined by the shape of the posterior calcaneal articular surface was directed less anteroposteriorly in humans than in apes, in contrast to the fact that the subtalar axis is more anteroposteriorly oriented in humans. It was also demonstrated that the navicular articular surface in humans was more plantarly oriented and axially twisted. These specialized features of the human talus seem to be functionally linked to obligate bipedal locomotion. The talar morphological differences among the great apes were prominent in the mediolateral and rotational orientations of the navicular articular surfaces, possibly reflecting the degree of arboreality among the great apes.     

Identification of a cyclooxygenase gene from the red alga Gracilaria vermiculophylla and bioconversion of arachidonic acid to PGF(2α) in engineered Escherichia coli

Prostaglandins (PGs) are important local messenger molecules in many tissues and organs of animals including human. For applications in medicine and animal care, PGs are mostly purified from animal tissues or chemically synthesized. To generate a clean, reliable, and inexpensive source for PGs, we have now engineered expression of a suitable cyclooxygenase gene in Escherichia coli and achieved production levels of up to 2.7 mg l⁻¹ PGF_2α. The cyclooxygenase gene cloned from the red alga Gracilaria vermiculophylla appears to be fully functional without any eukaryotic modifications in E. coli. A crude extract of the recombinant E. coli cells is able to convert in vitro the substrate arachidonic acid (AA) to PGF_2α. Furthermore, these E. coli cells produced PGF_2α in a medium supplemented with AA and secreted the PGF_2α product. To our knowledge, this is the first report of the functional expression of a cyclooxygenase gene and concomitant production of PGF_2α in E. coli. The successful microbial synthesis of PGs with reliable yields promises a novel pharmaceutical tool to produce PGF_2α at significantly reduced prices and greater purity.     

Certification of the critical importance of L-3-(2-naphthyl)alanine at position 3 of a specific CXCR4 inhibitor, T140, leads to an exploratory performance of its downsizing study

We have previously found that a 14-amino acid residue-peptide, T140, inhibits infection of target cells by T cell line-tropic HIV-1 (X4-HIV-1) through its specific binding to a chemokine receptor, CXCR4. Here, the importance of an L-3-(2-naphthyl)alanine (Nal) residue at position 3 in T140 for high anti-HIV activity and inhibitory activity against Ca(2+) mobilization induced by stromal cell-derived factor (SDF)-1alpha-stimulation through CXCR4 has initially been shown by the synthesis and biological evaluation of several analogues, where Nal(3) is substituted by diverse aromatic amino acids. Next, the order of the N-terminal 3 residues (Arg(1)-Arg(2)-Nal(3)) has been proved to be important from the structure--activity relationship (SAR) study shuffling these residues. Based on these results, we have found 10-residue peptides possessing modest anti-HIV activity by systematic antiviral evaluation of a series of synthetic, shortened analogues of T140.     

Molecular mechanisms of the synergistic induction of ornithine decarboxylase by asparagine and glucagon in primary cultured hepatocytes

In primary cultures of adult rat hepatocytes maintained in a salts/glucose medium, a more than 100-fold increase in ornithine decarboxylase (EC 4.1.1.17) activity was caused by asparagine and glucagon in a synergistic manner. The synthesis rate of ornithine decarboxylase was determined by [35S]methionine incorporation into the enzyme protein, and the amount of ornithine decarboxylase-mRNA was measured by hybridization with a cloned rat liver ornithine decarboxylase-cDNA. The synthesis rate of ornithine decarboxylase was stimulated more than 20-fold by asparagine and glucagon together, but the amount of ornithine decarboxylase-mRNA was increased only 3-4-fold, indicating that translational stimulation was involved in the induction process. Asparagine alone stimulated the synthesis of ornithine decarboxylase without substantial effect on the amount of ornithine decarboxylase-mRNA, whereas glucagon alone increased the amount of ornithine decarboxylase-mRNA about 3-fold without a detectable change in either enzyme activity or enzyme synthesis. Asparagine, at least in part, also suppressed degradation of ornithine decarboxylase.     

Composition of cardiolipin molecular species in Escherichia coli.

The composition of the molecular species of acidic phospholipids in Escherichia coli B during the late exponential growth phase at 37 degrees C was determined. Two phosphatidyl groups of cardiolipin, the 3-(3-sn-phosphatidyl) and 1-(3-sn-phosphatidyl) moieties of cardiolipin, were isolated by limited hydrolysis with phospholipase C. No significant difference in the composition of the molecular species was found between the 3-(3-sn-phosphatidyl) and 1-(3-sn-phosphatidyl) moieties. On the other hand, the composition of the molecular species of phosphatidylglycerol was different from that of cardiolipin. Phosphatidylglycerol contained more of the 1-palmitoyl 2-cis-9,10-methylenehexadecanoyl and 1-palmitoyl 2-cis-11,12-methyleneoctadecanoyl species than did cardiolipin. The difference in the composition of the molecular species between cardiolipin and phosphatidylglycerol may depend on the difference in the turnover rates of both phospholipids.     

Analyses of ornithine decarboxylase antizyme mRNA with a cDNA cloned from rat liver

Ornithine decarboxylase antizyme is a unique inhibitory protein induced by polyamines and involved in the regulation of ornithine decarboxylase. A cDNA was isolated from a rat liver cDNA library by the screening with monoclonal antibodies to rat liver antizyme as probes. The expression products of the cDNA in bacterial systems inhibited rat ornithine decarboxylase activity in a manner characteristic of antizyme and rabbit antisera raised against its direct expression product reacted to rat liver antizyme, confirming the authenticity of the cDNA. On RNA blot analysis with the cDNA probe, an antizyme mRNA band of 1.3 kb was detected in rat tissues. Antizyme mRNA did not increase upon administration of putrescine, an inducer of antizyme, and its half-life after actinomycin D treatment was as long as 12 h in rat liver, suggesting that antizyme mRNA is constitutively expressed and antizyme synthesis is regulated at the translational level. Similar-sized mRNAs hybridizable to the cDNA were also found in various mammalian and non-mammalian vertebrate tissues under physiological conditions. In addition, chicken and frog antizymes showed immunocrossreactivity with rat antizyme. The ubiquitous presence and the evolutionally conserved structure of antizyme in vertebrate tissues suggest that it has an important function.