Bacteriophages of l-Glutamic Acid-Producing Bacteria

The growth characteristics of phages were investigated with the four phages, active on Brevibacterium lactofermentum, which were selected from the respective serological groups, namely, P465 (group I), P468II (group II), Ap85III (group III) and P4 (group IV).

The adsorption rate of the phages, P465 and P468II, on the host bacteria was low, whereas that of the phages, Ap85III and P4, was higher. The adsorption rate constants for the four phages were respectively calculated at 2.02 × 10^?10, 1.87 × 10^?10, 4.32 × 10^?10 and 3.15 × 10^?10 cm³ per minute, at 30°C in G₅B₂ medium. With reference to the ionic environment for adsorption, the phages, P465 and Ap85III, specifically required either for Ca⁺⁺ or Mg⁺⁺; the phage P468II, for both; and the phage P4, for neither.

The growth characteristics of these phages were examined by the one-step growth experiment. The latent periods of the phages were 50, 53, 57 and 47 minutes, respectively; and the corresponding average burst sizes were about 98, 31, 145 and 126. The growth of the phage P4 was completely suppressed at above 34°C, although the host bacteria and the other three phages were capable of the full growth at that temperature.     

Single-laboratory validation for the determination of caffeic acid and seven caffeoylquinic acids in sweet potato leaves

A single-laboratory validation study was conducted on an HPLC method for the detection and quantification of caffeic acid (CA) and seven species of caffeoylquinic acids (CQAs) in lyophilized sweet potato leaves. The procedure for extraction of the analytes from the matrix and the HPLC conditions for the efficient separation of CA and CQAs were optimized. In the proposed method, a relative response factor to one of the CQAs (5-CQA) was used to quantify the others. The method performed well in terms of precision when carried out on five different days and demonstrated Horwitz ratio (HorRat) scores ranging from 0.5 to 1.0 for all analytes, which were well within the limits of performance acceptability. Accuracy testing at three levels showed an overall recovery of 94% when duplicated on five different days. Moreover, a stability study revealed that all analytes in both standard solution and sample extract were stable for 28?days.     

Bacteriophages of l-Glutamic Acid-Producing Bacteria

Some of the characteristics of the four phages, P 465 (group I), P 468 II (group II), Ap 85 III (group III) and P 4 (group IV), of Brevibacterium lacto fermentum, i.e., stability in salt solution, thermal and pH stabilities, inactivation by ultraviolet irradiation, stability on air-drying, and lytic characteristics of infected cells, are descrided. We have observed differences in these physico-chemical characteristics among these four phages selected from the respective serological groups. An addition of protein such as casein to diluent at a 0.01 ~ 1% concentration increased remarkably the stabilities on shaking of these phages except Ap 85 III. There were great differences in stabilities on air-drying among the four phages. Phage P 61 (gropu I) which was isolated from the first abnormal fermentation broth showed the lowest stability in the dry state, while phages, P 468 II and Ap 85 III, were very stable and were not inactivated by the relative humidity of 10% at 30°C, for the period of five months. In general, Ар-series phages isolated from air-borne particles at fermentation factory can be distinguished from the other P-series phages which were isolated from abnormal fermentation broth, by their considerably high resistance to drying.     

Gas Chromatographic Determination of a New Fungicide,Procymidone (Sumilex®) in Technical Preparation and Formulation

Monoclonal antibodies (MoAbs) reacting with human G-CSF and /or its muteins were established by cell fusion between P3.X63/Ag8.U1 myeloma cells and spleen cells from BALB/C mice immunized with recombinant human intact G-CSF or its mutein, designated ND28. Two MoAbs reacted with intact G-CSF and all kinds of muteins tested, designated KM341 and KM342, and two MoAbs specific for intact G-CSF, designated KM340 and KM343, were obtained from the mice immunized with recombinant human intact G-CSF.

The sera from the mice immunized with a mutein of G-CSF, ND28, reacted with intact G-CSF and all muteins tested. Two MoAbs specific for ND28, designated KM498 and KM511, were obtained from these mice. These MoAbs seem to recognize the sequence of a few amino acids that is peculiar for ND28. However, the epitopes recognized by KM498 and KM511 were maybe subtly different, because KM498 and KM511 could not completely inhibit each other.

Human G-CSF and/or its muteins could be measured by sandwich ELISA using these MoAbs with suitable combinations. The immuno-affinity column using KM342 or KM498 adsorbed G-CSFs or specifically ND28, previously. By elution with 0.15m NH₄OH, the G-CSFs or ND28 were eluted with a high recovery.     

The effects of an arabinogalactan-protein from the white-skinned sweet potato (Ipomoea batatas L.) on blood glucose in spontaneous diabetic mice

We examined the effects of an arabinogalactanprotein (WSSP-AGP) from Ipomoea batatas L. on hyperglycemia in db/db mice. An oral glucose tolerance test indicated significantly decreased plasma glucose levels by WSSP-AGP. Additionally, an insulin tolerance test found improvement in insulin sensitivity due to treatment with WSSP-AGP. This suggests that amelioration of insulin resistance by WSSP-AGP causes to lead its hypoglycemic effects.     

Crystal structures of protein glutaminase and its pro forms converted into enzyme-substrate complex

Protein glutaminase, which converts a protein glutamine residue to a glutamate residue, is expected to be useful as a new food-processing enzyme. The crystal structures of the mature and pro forms of the enzyme were refined at 1.15 and 1.73 ? resolution, respectively. The overall structure of the mature enzyme has a weak homology to the core domain of human transglutaminase-2. The catalytic triad (Cys-His-Asp) common to transglutaminases and cysteine proteases is located in the bottom of the active site pocket. The structure of the recombinant pro form shows that a short loop between S2 and S3 in the proregion covers and interacts with the active site of the mature region, mimicking the protein substrate of the enzyme. Ala-47 is located just above the pocket of the active site. Two mutant structures (A47Q-1 and A47Q-2) refined at 1.5 ? resolution were found to correspond to the enzyme-substrate complex and an S-acyl intermediate. Based on these structures, the catalytic mechanism of protein glutaminase is proposed.     

Invariant NKT cells biased for IL-5 production act as crucial regulators of inflammation

Although invariant NKT (iNKT) cells play a regulatory role in the pathogenesis of autoimmune diseases and allergy, an initial trigger for their regulatory responses remains elusive. In this study, we report that a proportion of human CD4+ iNKT cell clones produce enormous amounts of IL-5 and IL-13 when cocultured with CD1d+ APC in the presence of IL-2. Such IL-5 bias was never observed when we stimulated the same clones with alpha-galactosylceramide or anti-CD3 Ab. Suboptimal TCR stimulation by plate-bound anti-CD3 Ab was found to mimic the effect of CD1d+ APC, indicating the role of TCR signaling for selective induction of IL-5. Interestingly, DNA microarray analysis identified IL-5 and IL-13 as the most highly up-regulated genes, whereas other cytokines produced by iNKT cells, such as IL-4 and IL-10, were not significantly induced. Moreover, iNKT cells from BALB/c mice showed similar IL-5 responses after stimulation with IL-2 ex vivo or in vivo. The iNKT cell subset producing IL-5 and IL-13 could play a major role in the development of allergic disease or asthma and also in the immune regulation of Th1 inflammation.     

Sulfated glycosaminoglycans are necessary for Nodal signal transmission from the node to the left lateral plate in the mouse embryo

Situs-specific organogenesis in the mouse results from leftward fluid flow in the node cavity and subsequent left-sided expression of Nodal in the lateral plate mesoderm (LPM). Nodal expression at the node is essential for the subsequent asymmetric Nodal expression in the left LPM, but the precise role of Nodal produced at the node has remained unknown. We have now investigated how the Nodal signal is transferred from the node to the LPM. Externally supplied Nodal protein failed to signal to the LPM, suggesting that the Nodal signal is transferred to the LPM via an internal route rather than an external one. Transgenic rescue experiments showed that the Nodal co-receptor Cryptic (Cfc1) is required only in the LPM, not at the node, for asymmetric Nodal expression in the LPM, indicating that the Nodal signal is not relayed indirectly between the node and LPM. Nodal interacts in vitro with sulfated glycosaminoglycans (GAGs), which are specifically localized to the basement membrane-like structure between the node and LPM in the mouse embryo. Inhibition of sulfated GAG biosynthesis prevents Nodal expression in the LPM. These data suggest that Nodal produced at the node might travel directly to the LPM via interaction with sulfated GAGs.