l-Glutamic Acid Fermentation

A study was made on the differences between Brevibacterium thiogenitalis No. 653 and its oleic acid-requiring mutant D-248 in some physiological characteristics.

The most important difference of the characteristics was found in their intracellular fatty acid contents. Namely, the cellular oleic acid content of D-248 was scarcely affected by biotin but limited by the oleic acid which was added to the medium.

On the other hand, various enzyme activities and rates of oxygen uptake for several organic acids were found to be slightly different between the two strains.

These observations suggest that oleic acid has an important role for the production of l-glutamic acid.

The effect of biotin and oleic acid on the cellular fatty acid contents, and the relation between the cellular fatty acid contents and the productivity of l-glutamic acid were investigated using Brevibacterium thiogenitalis No. 653 and its oleic acid-requiring mutant, D-248.

While the synthesis of palmitic acid in D-248 was stimulated by biotin and competitively reversed by oleic acid added to the culture medium, the level of cellular oleic acid was scarcely affected by biotin but regulated by oleic acid in the medium.

For the productivity of L-glutamic acid, the most important factor was the level of cellular oleic acid, and the effect of cellular palmitic acid was considerably weak. This relation was subjected to a figuration and able to be expressed on the whole as one exponential-like curve. An amount of over 70 per cent of cellular fatty acids was distributed in the phospholipid fraction and its fatty acid composition was almost the same as that of whole cells.     

Forced expression of FLO11 confers pellicle-forming ability and furfural tolerance on Saccharomyces cerevisiae in ethanol production

We constructed a plasmid that expresses FLO11 encoding a cell surface glycoprotein of Saccharomyces cerevisiae under the control of a constitutive promoter. This plasmid conferred pellicle-forming ability on the non-pellicle-forming industrial strain of S. cerevisiae at the air–liquid interface of the glucose-containing liquid medium. The induced pellicle-forming cells exhibited tolerance to furfural, which is a key toxin in lignocellulosic hydrolysates, in ethanol production.     

MACROSCOPIC OBSERVATIONS OF THE FACIAL MUSCLES IN THE BAIKAL SEAL (PHOCA SIBIRICA)

This study was performed to determine if, as expected, the enlarged eye of the Baikal seal ( Phoca sibirica ) has an influence on the form and function of the skull and facial muscles. Macroscopic observation of these muscles demonstrated that the M. orbicularis oculi expands around the palpebral fissure and that some facial muscles attach and insert in the M. orbicularis oculi , possibly supporting M. orbicularis oculi function. We suggest that these muscles move the eye and palpebral area and constitute a morphological and synergistic facial muscle complex system. Further, the development of the M. rectus lateralis around the sclera of the eye indicates that this muscle is also involved in eye movement.     

Standardized data and relationship between bone growth and bone metabolism in female G?ttingen minipigs.

Minipigs have been studied as a model of osteoporosis. However, little information is available regarding their bone physiology. We established standardized bone data and investigated the relationship between bone growth and bone metabolism in female minipigs. Blood and urine samples were obtained from 53 female G?ttingen minipigs, 3-76 months of age, for measurement of bone biomarkers (i.e., BAP, OC, NTX, and DPD). The lumbar vertebra and femur were excised to determine the growth plate condition, bone length, bone mineral content (BMC), and bone mineral density (BMD). High levels of bone biomarkers were observed during the initial period after birth, decreasing thereafter with age. Bone biomarkers were confirmed to be highly correlated with age (R(2) > 0.7). The growth plates of the lumbar vertebra and the femur began to close at 21 and 25 months of age, respectively, and closed completely at 42 months of age. Bone length increased rapidly before growth plate closure, and reached a peak at 21 and 28 months of age in the lumbar vertebra and the femur, respectively. The levels of BMC and BMD increased rapidly before growth plate closure, and continued to increase slowly until 76 months of age. A high negative correlation (-0.855 < r < -0.711, p<0.001) was confirmed between the bone biomarkers and the bone measurement data. These results indicate that the bone turnover velocity is consistent with the bone growth velocity in female G?ttingen minipigs.     

Response of hepatic function to hepatic copper deposition in rats fed a diet containing copper

Fischer rats were a fed diet supplied with copper chloride (150–600 ppm) for 60 d from weaning. Serum (glutamic-oxaloacetic transaminase (GOT) and glutamic-pyruvic transaminase (GPT) activities were increased with the increase of Cu concentration in the diet. Biliary excretion of Cu was related to the dietary Cu level. Depositions of hepatic and renal Cu were also related to the dietary Cu level in a dose-dependent manner. In particular, hepatic (155.2±13.3 μg/g) and renal (44.9±4.4 μg/g) Cu concentrations increased abruptly in the Cu-600 ppm group. In the liver, about 60% of Cu was distributed in the soluble fraction (100,000 g supernatant). In the Cu-600 ppm group, 25% of cystosolic Cu was bound to metallothionein (MT). Our results suggest that chronic exposure to Cu appears to have a deleterious effect on the hepatic function, and further, that even in rats with normal biliary Cu excretion, clearance of Cu from the liver may be marginal when dietary Cu is near the 600-ppm level. Although Cu is an essential nutrient, an overload of Cu should be avoided.     

But1 and But2 proteins bind to Uba3, a catalytic subunit of E1 for neddylation, in fission yeast

NEDD8/Rub1 is the most homologous protein to ubiquitin among the ubiquitin-like proteins, and it is covalently linked to target proteins via the C-terminal glycine residue in a manner analogous to ubiquitylation. However, the mechanism(s) involved in the regulation of the NEDD8 ligation pathway remains elusive. Using the two-hybrid system, we isolated novel genes from the Schizosaccharomyces pombe cDNA library whose products bind to Uba3, which is a catalytic protein for E1-like activity of the NEDD8 pathway. We designated these genes but1(+) and but2(+) (for proteins that bind to Uba three). But1 is a nuclear protein and its overexpression caused cell elongation, which is a common phenotype of the NEDD8 pathway defective mutant in S. pombe. Furthermore, overexpression of but1(+) in ned8-temperature sensitive mutant had a deleterious effect even under permissive temperatures. Our results suggest that But1 may have an inhibitory role in the NEDD8 pathway.     

Non-additive effects of leaf litter and insect frass mixture on decomposition processes

Although there is a growing body of evidence that herbivorous insects have a significant impact on decomposition and soil nutrient dynamics through frass excretion, how mixtures of leaf litter and insect frass influence such ecosystem processes remains poorly understood. We examined the effects of mixing of leaf litter and insect frass on decomposition and soil nutrient availability, using a study system consisting of a willow, Salix gilgiana Seemen, and a herbivorous insect, Parasa consocia Walker. The chemical characteristics of insect frass differed from those of leaf litter. In particular, frass had a 42-fold higher level of ammonium–nitrogen (NH₄ ⁺–N) than litter. Incubation experiments showed that the frass was decomposed and immobilized with respect to N more rapidly than the litter. Furthermore, litter and frass mixtures showed non-additive enhancement of decomposition and reduction of NH₄ ⁺–N, depending on the litter–frass mixing ratio. These indicate that, while insect frass generally accelerated decomposition, the effect of frass on soil nutrient availability was dependent largely on the relative amounts of litter and frass.     

CD4+ T cell clones specific for the human p97 melanoma-associated antigen can eradicate pulmonary metastases from a murine tumor expressing the p97 antigen.

p97 is a human tumor-associated Ag present on most melanoma cells that represents a possible target for immunologic attack. To evaluate the capacity of T cells reactive with this protein to promote elimination of melanoma cells expressing p97, a murine model was developed by transfecting a C3H/HeN melanoma with the p97 cDNA, generating p97-specific CD4+ T cells by in vivo immunization of C3H/HeN mice with a vaccinia/p97 recombinant virus followed by in vitro cloning with soluble p97 protein, and determining whether these CD4+ T cells could mediate rejection of pulmonary metastases. Characterization of the T cell clones demonstrated the presence of both I-Ak and I-Ek-restricted clones, although the majority of clones recognized p97 in the context of I-Ek. Analysis of clonal specificity using truncated p97 proteins revealed that at least three epitopes were immunogenic, and further studies with overlapping 15-amino acid peptides from a region of the p97 molecule defined by these truncated proteins identified an immunodominant epitope responsible for the majority of the I-Ek response. The T cell clones were not capable of directly recognizing the p97-expressing melanoma cells but responded to the tumor if syngeneic APC were present to process the tumor-derived p97 Ag. The therapeutic efficacy of these CD4+ T cell clones was evaluated in an adoptive therapy model in which mice bearing metastatic pulmonary lesions were treated by i.v. administration of the p97-specific cells. Despite the inability of the CD4+ clones to directly respond to or lyse the tumor cells, the clones were effective in promoting tumor eradication. In vitro studies demonstrated that this may have reflected secretion of lymphokines that activated macrophages to lyse the tumor. The results suggest that noncytolytic p97-specific CD4+ T cell clones can be effective in therapy of pulmonary melanoma metastases. Moreover, if human T cells reactive with the p97 protein could be generated, the expression of this tumor-associated Ag in melanoma cells might be adequate for such T cells to mediate a therapeutic antitumor response.