Effect of low protein and low energy diet on physiological status and digestibility of F344 rats.

A long-term raising study was carried out on male F344/DuCrj rats with three low protein (Crude Protein (CP); 14.5, 11.5, 8.5%) and low energy (Digestible Energy (DE); 2.0 kcal/g) diets from 4 to 104 weeks of age. In rats fed the 8.5% CP diet, body weight and digestible crude protein (DCP) consumption at 10 weeks of age were lower (P < 0.05) but the body weight at 50 weeks of age was higher (P < 0.05) than in the other groups. In rats fed the 8.5% CP diet the crude fat digestibility was higher (P < 0.05), and the CP/nitrogen-corrected metabolizable energy (MEn) ratio was low. On the other hand, the mean survival time at 80 weeks of age was shorter in rats fed the 8.5% CP diet (P < 0.05).     

Inhibiting the Activity of CA1 Hippocampal Neurons Prevents the Recall of Contextual Fear Memory in Inducible ArchT Transgenic Mice

The optogenetic manipulation of light-activated ion-channels/pumps (i.e., opsins) can reversibly activate or suppress neuronal activity with precise temporal control. Therefore, optogenetic techniques hold great potential to establish causal relationships between specific neuronal circuits and their function in freely moving animals. Due to the critical role of the hippocampal CA1 region in memory function, we explored the possibility of targeting an inhibitory opsin, ArchT, to CA1 pyramidal neurons in mice. We established a transgenic mouse line in which tetracycline trans-activator induces ArchT expression. By crossing this line with a CaMKIIα-tTA transgenic line, the delivery of light via an implanted optrode inhibits the activity of excitatory CA1 neurons. We found that light delivery to the hippocampus inhibited the recall of a contextual fear memory. Our results demonstrate that this optogenetic mouse line can be used to investigate the neuronal circuits underlying behavior.     

Enhancement of Cellulose Degradation by Cattle Saliva

Saccharification of cellulose is a promising technique for producing alternative source of energy. However, the efficiency of conversion of cellulose into soluble sugar using any currently available methodology is too low for industrial application. Many additives, such as surfactants, have been shown to enhance the efficiency of cellulose-to-sugar conversion. In this study, we have examined first whether cattle saliva, as an additive, would enhance the cellulase-catalyzed hydrolysis of cellulose, and subsequently elucidated the mechanism by which cattle saliva enhanced this conversion. Although cattle saliva, by itself, did not degrade cellulose, it enhanced the cellulase-catalyzed degradation of cellulose. Thus, the amount of reducing sugar produced increased approximately 2.9-fold by the addition of cattle saliva. We also found that non-enzymatic proteins, which were present in cattle saliva, were responsible for causing the enhancement effect. Third, the mechanism of cattle saliva mediated enhancement of cellulase activity was probably similar to that of the canonical surfactants. Cattle saliva is available in large amounts easily and cheaply, and it can be used without further purification. Thus, cattle saliva could be a promising additive for efficient saccharification of cellulose on an industrial scale.     

The Catalytic Domain of Topological Knot tRNA Methyltransferase (TrmH) Discriminates between Substrate tRNA and Nonsubstrate tRNA via an Induced-fit Process

A conserved guanosine at position 18 (G18) in the D-loop of tRNAs is often modified to 2′-O-methylguanosine (Gm). Formation of Gm18 in eubacterial tRNA is catalyzed by tRNA (Gm18) methyltransferase (TrmH). TrmH enzymes can be divided into two types based on their substrate tRNA specificity. Type I TrmH, including Thermus thermophilus TrmH, can modify all tRNA species, whereas type II TrmH, for example Escherichia coli TrmH, modifies only a subset of tRNA species. Our previous crystal study showed that T. thermophilus TrmH is a class IV S-adenosyl-l-methionine-dependent methyltransferase, which maintains a topological knot structure in the catalytic domain. Because TrmH enzymes have short stretches at the N and C termini instead of a clear RNA binding domain, these stretches are believed to be involved in tRNA recognition. In this study, we demonstrate by site-directed mutagenesis that both N- and C-terminal regions function in tRNA binding. However, in vitro and in vivo chimera protein studies, in which four chimeric proteins of type I and II TrmHs were used, demonstrated that the catalytic domain discriminates substrate tRNAs from nonsubstrate tRNAs. Thus, the N- and C-terminal regions do not function in the substrate tRNA discrimination process. Pre-steady state analysis of complex formation between mutant TrmH proteins and tRNA by stopped-flow fluorescence measurement revealed that the C-terminal region works in the initial binding process, in which nonsubstrate tRNA is not excluded, and that structural movement of the motif 2 region of the catalytic domain in an induced-fit process is involved in substrate tRNA discrimination.     

Assimilation of Formaldehyde and other C1Compounds by Gliocladium deliquescens and Paecilomyces varioti

Two fungi were isolated from soil which grew on 0.1~0.2% formaldehyde as the sole carbon source, and identified as Gliocladium deliquescens and Paecilomyces varioti. Both the strains could grow on 5% methanol and 5% Na-formate, while the former could grow even on 7% methanol. Metabolic pathways were traced through two dimensional paper chromatography and autoradiographic techniques using ¹⁴C-formaldehyde, ¹⁴C-methanol or ¹⁴C-CO₂ as substrates.

The intracellular metabolites were persued and their quantitative variation with time was measured. Along with the fact that serine and malate appeared in the earlier time, then appeared organic acids and amino acids belonging to TCA cycle, and the fact that hydroxy-pyruvate reductase and phosphoenolpyruvate carboxylase activities were much stronger in methanol culture than in ethanol culture, it was concluded that the two fungi followed the serine pathway in assimilating C₁-compounds. Oxidation enzymes of methanol and formaldehyde were also studied, and an oxidizing system was found besides usual NAD linked methanol or formaldehyde dehydrogenases.     

Trigonelline Content in Coffee Beans and the Thermal Conversion of Trigonelline into Nicotinic Acid during the Roasting of Coffee Beans

The trigonelline content in coffee was determined by the microbiological assay method after demethylating the compound. The content was proved to be extremely high (up to 1 % on the wet basis). When trigonelline was heated to above 180°C, it was converted into nicotinic acid. Although the conversion rate was low, a nutritionally significant amount of nicotinic acid was formed during roasting coffee beans because of the high content of trigonelline in coffee beans. The optimum heating condition for nicotinic acid formation was found at 220°C for 20 min.     

Some Enzymatic Properties and Substrate Specificities of Peptidoglutaminase-I and II

Some enzymatic properties and substrate specificities of the two peptidoglutaminases (peptidoglutaminase-I (PGase-I) and II (PGase-II) isolated from Bacillus circulans were investigated.

The both resembled each other with respect to optimum pH and temperature for their activities, susceptibility to various reagents and effects of many metal ions. But, the pH-rate profile of PGase-II was more broad than that of PGase-I. Thermal stability of PGase-I was better than that of PGase-II at a degree of about 5°C.

However, they were definitively different each other with respect to their substrate specificities. PGase-I specifically deamidated the γ-amide of L-glutamine residing at the carboxyl terminal on peptides, and was inactive to glutamine derivative substituted at both α-amino and α-carboxyl groups. On the other hand, the best substrate for PGase-II was tri-peptides, X-Gln-Y, where X was carbobenzoxy-,t-amyloxycarbonxy- or amino acid residues, and Y was amino acids. Though L-glutamine presented in polypeptide chains composed of more than four amino acid residues was a poor substrate, two L-glutamines in oxidized insulin A chain were well attacked by the enzyme.

The both PGase were inactive to asparagine and asparaginyl peptides.     

Lysis of Pediococcus and Brevibacterium Cells by Cell-Wall Lytic Enzymes,L3, L11, ALE and Lysozyme

Several species belonging to the genera Pediococcus and Brevibacterium, which have resistant cell walls against the usual cell-disrupting methods, were effectively attacked by new cell-wall lytic enzymes, L₃, L₁₁ and ALE which were obtained from a Streptomyces sp., a Flavobacterium sp., and a Staphylococcus sp., respectively. Among them, the L₃ enzyme was mostly effective to all pediococcal and brevibacterial strains.     

P-Factor,a New Peptidic Growth Factor for Pediococcus soyae

The P-factor a new growth factor for Ped. soyae, is found in partial acid and enzymic hydrolysates of Hammersten’s milk casein, and is therefore, supposed to be a kind of peptide. The well-known peptidic growth promoting substance for Lact. casei, strepogenin and its relating substances, are quite ineffective to the author’s organism. P-factor exists along with the S-factor which is another growth promoting-factor for this lactic acid bacterium, in soy mash juice and in the water extract of Aspergillus sojae mycelium. The former substance is also found in various kinds of commercial peptones. The factor was not substitutable with all of the 19 synthetic peptides tested. S-factor is effective, only in the presence of the P-factor.