Further Studies on Production of Chloramphenicol Analogs (Corynecins) from n-Alkanes

Further studies on culture condition of Corynebacterium hydrocarboclastus were carried out for more efficient production of corynecins (chloramphenicol analogs). The productivity was affected greatly by the concentration of phosphate and stimulated effectively by organic nutrients, particularly by yeast extract. The most effective production was obtained in the presence of 8 g of yeast extract and 0.2 g of K₂HPO₄ per one liter of chemically defined medium. The maximum amount was 1.42 g equivalent of l-base (free base of chloramphenicol) per liter of the broth culture, which corresponded to 4-fold of that reported in the previous paper.

Among fractions of n-alkanes, n-heptadecane was the best carbon source for the production. The proportion among corynecins homologs varied depending on carbon number of n-alkane used; odd number alkanes gave larger amounts of corynecin II (propionyl derivative), suggesting that carbon flow in the bacterial cells functioned significantly in changing the ratios of corynecin homologs.     

Effects of ingestion of difructose anhydride III (DFA III) and the DFA III-assimilating bacterium Ruminococcus productus on rat intestine

We have isolated a difructose anhydride III (DFA III)-assimilating bacterium, Ruminococcus productus AHU1760, from human. After an acclimation period of 1 week, male Sprague-Dawley rats (5 weeks old) were divided into four groups (control diet, R. productus diet, DFA III diet, and R. productus + DFA III diet; n = 8) and fed the assigned test diets for 2 weeks. The viable count of administered R. productus was 4.9 x 10(7) CFU/d in R. productus-fed rats and 4.7 x 10(7) CFU/d in R. productus + DFA III-fed rats. Survival in cecal content of this strain was confirmed by randomly amplified polymorphic DNA. The ratio of secondary bile acids in feces in R. productus + DFA III-fed rats decreased the same as that in rats fed only DFA III. The viable count of lactobacilli and bifidobacteria, known as beneficial bacteria, increased more in R. productus + DFA III-fed rats than in control or R. productus-fed rats. A combination of R. productus and DFA III might improve the balance of intestinal microbiota to a healthier condition.     

Prostaglandin F2alpha formation from prostaglandin H2 by prostaglandin F synthase (PGFS): crystal structure of PGFS containing bimatoprost

Prostaglandin H(2) (PGH(2)) formed from arachidonic acid is an unstable intermediate and is efficiently converted into more stable arachidonate metabolites by the action of enzymes. Prostaglandin F synthase (PGFS) has dual catalytic activities: formation of PGF(2)(alpha) from PGH(2) by the PGH(2) 9,11-endoperoxide reductase activity and 9alpha,11beta-PGF(2) (PGF(2)(alphabeta)) from PGD(2) by the PGD(2) 11-ketoreductase activity in the presence of NADPH. Bimatoprost (BMP), which is a highly effective ocular hypotensive agent, is a PGF(2)(alpha) analogue that inhibits both the PGD(2) 11-ketoreductase and PGH(2) 9,11-endoperoxide reductase activities of PGFS. To examine the catalytic mechanism of PGH(2) 9,11-endoperoxide reductase, a crystal structure of PGFS[NADPH + BMP] has been determined at 2.0 A resolution. BMP binds near the PGD(2) binding site, but the alpha- and omega-chains of BMP are locate on the omega- and alpha-chains of PGD(2), respectively. Consequently, the bound BMP and PGD(2) direct their opposite faces of the cyclopentane moieties toward the nicotinamide ring of the bound NADP. The alpha- and omega-chains of BMP are involved in H-bonding with protein residues, while the cyclopentane moiety is surrounded by water molecules and is not directly attached to either the protein or the bound NADPH, indicating that the cyclopentane moiety is movable in the active site. From the complex structure, two model structures of PGFS containing PGF(2)(alpha) and PGH(2) were built. On the basis of the model structures and inhibition data, a putative catalytic mechanism of PGH(2) 9,11-endoperoxide reductase of PGFS is proposed. Formation of PGF(2)(alpha) from PGH(2) most likely involves a direct hydride transfer from the bound NADPH to the endoperoxide of PGH(2) without the participation of specific amino acid residues.     

Serum levels of trace elements and iron-deficiency anemia in adult Vietnamese

This study was aimed at assessing the serum levels of vitamin A, copper, zinc, selenium, and iron among adult Vietnamese with and without iron-deficiency anemia. Blood was collected from adult Vietnamese living in the midland of northern Vietnam. One hundred twenty-three subjects in the age range 20–60 yr were included in the study. Anemia, where the concentration of hemoglobin in whole blood is less than 120 g/L in females and 130 g/L in males, was found in 30% (37/123) of the study population. The levels of vitamin A and selenium in the sera of anemic subjects (n=37) were significantly lower than that in nonanemic group (n=86). On the other hand, no significant differences were observed in the concentrations of copper and zinc between the two groups. This study was the first to show serum levels of trace elements in adult Vietnamese, providing useful baseline information for further studies.     

A catalytic mechanism that explains a low catalytic activity of serine dehydratase like-1 from human cancer cells: crystal structure and site-directed mutagenesis studies

SDH (l-serine dehydratase, EC 4.3.1.17) is a pyridoxal-5'-phosphate (PLP)-dependent enzyme that catalyzes dehydration of l-Ser/Thr to yield pyruvate/ketobutyrate and ammonia. A SDH isoform (cSDH) found in human cancer cell lines has relatively low catalytic activity in comparison with the liver enzyme (hSDH). The crystal structure of cSDH has been determined at 2.8 angstroms resolution. A PLP is covalently attached to K48 by Schiff-base linkage in the active site. The ring nitrogen of PLP is involved in a H-bonding with C309, but is apparently not protonated. Twenty-three amino residues that compose the active site surfaces were identified. The human and rat liver enzymes (hSDH and rSDH) have the same residues, while residues G72, A172, and S228 in cSDH are replaced with A66, S166, and A222, respectively, in hSDH. These residues in hSDH and cSDH were mutated to make complementary pairs of mutated enzymes, and their kinetic parameters were determined. C303 of hSDH and C309 of cSDH which are H-bonding partner of the ring nitrogen of PLP were mutated to alanine and their kinetic parameters were also determined. The crystal structures and the mutation data suggest that having a glycine at residue 72 of cSDH is the major reason for the reduction of catalytic activity of cSDH. Changing alanine to glycine at residue 72 increases the flexibility of the substrate binding-loop (71S(G/A)GN74), so that the bound substrate and PLP are not pushed deep into the active cleft. Consequently, the proton transfer rate from S(G) of C309 to N1 of the bound PLP is decreased, which determines the rate of catalytic reaction.     

Localization and hormonal control of serine dehydratase during metabolic acidosis differ markedly from those of phosphoenolpyruvate carboxykinase in rat kidney

Serine dehydratase (SDH) is abundant in the rat liver but scarce in the kidney. When administrated with dexamethasone, the renal SDH activity was augmented 20-fold, whereas the hepatic SDH activity was affected little. In situ hybridization and immunohistochemistry revealed that SDH was localized to the proximal straight tubule of the nephron. To address the role of this hormone, rats were made acidotic by gavage of NH(4)Cl. Twenty-two hours later, the SDH activity was increased three-fold along with a six-fold increment in the phosphoenolpyruvate carboxykinase (PEPCK) activity, a rate-limiting enzyme of gluconeogenesis. PEPCK, which is localized to the proximal tubules under the normal condition, spreads throughout the entire cortex to the outer medullary rays by acidosis, whereas SDH does not change regardless of treatment with dexamethasone or NH(4)Cl. When NH(4)Cl was given to adrenalectomized rats, in contrast to the SDH activity no longer increasing, the PEPCK activity responded to acidosis to the same extent as in the intact rats. A simultaneous administration of dexamethasone and NH(4)Cl into the adrenalectomized rats fully restored the SDH activity, demonstrating that the rise in the SDH activity during acidosis is primarily controlled by glucocorticoids. The present findings clearly indicate that the localization of SDH and its hormonal regulation during acidosis are strikingly different from those of PEPCK.     

Strain-specific T-cell suppression and protective immunity in patients with chronic hepatitis C virus infection

Hepatitis C virus (HCV) frequently persists with an apparently ineffective antiviral T-cell response. We hypothesized that some patients may be exposed to multiple HCV subtypes and that strain-specific T cells could contribute to the viral dynamics in this setting. To test this hypothesis, CD4 T-cell responses to three genotype 1a-derived HCV antigens and HCV antibody serotype were examined in chronically HCV infected (genotypes 1a, 1b, 2, 3, and 4) and spontaneously HCV recovered subjects. Consistent with multiple HCV exposure, 63% of patients infected with genotypes 2 to 4 (genotypes 2-4) and 36% of those infected with genotype 1b displayed CD4 T-cell responses to 1a-derived HCV antigens, while 29% of genotype 2-4-infected patients showed serotype responses to genotype 1. Detection of 1a-specific T cells in patients without active 1a infection suggested prior self-limited 1a infection with T-cell-mediated protection from 1a but not from non-1a viruses. Remarkably, CD4 T-cell responses to 1a-derived HCV antigens were weakest in patients with homologous 1a infection and greater in non-1a-infected patients: proportions of patients responding were 19% (1a), 36% (1b), and 63% (2-4) (P = 0.0006). Increased 1a-specific CD4 T-cell responsiveness in non-1a-infected patients was not due to increased immunogenicity or cross-reactivity of non-1a viruses but directly related to sequence divergence. We conclude that the T-cell response to the circulating virus is either suppressed or not induced in a strain-specific manner in chronically HCV infected patients and that, despite their ability to clear one HCV strain, patients may be reinfected with a heterologous strain that can then persist. These findings provide new insights into host-virus interactions in HCV infection that have implications for vaccine development.     

Density-mediated indirect effects of a common prey tadpole on interaction between two predatory bugs: <Emphasis Type="Italic">Kirkaldyia deyrolli</Emphasis> and <Emphasis Type="Italic">Laccotrephes japonensis</Emphasis>

This paper suggests that the nymphs of a specialist predator, Kirkaldyia (=Lethocerus) deyrolli (Belostomatidae: Heteroptera), are indirectly affected by their tadpoles’ prey. K. deyrolli nymphs and the predator Laccotrephes japonensis (Nepidae: Heteroptera) adults coexist in rice paddy fields. It was predicted that the difference in tadpole density may influence the K. deyrolli nymph survival rate. We first compared survival rates of the first instar nymphs of K. deyrolli in June (high tadpole density period) and July (low tadpole density period). Secondly, we investigated the survival rate of K. deyrolli nymphs at different tadpole densities and under the presence or absence of L. japonensis adults to examine whether higher tadpole density moderates predation pressure from L. japonensis adults to K. deyrolli nymphs, e.g., density-mediated indirect effects. As a result of the comparison, the survival rate of K. deyrolli nymphs in June was higher than that in July. For the field experiment, the slopes between the survival rate of K. deyrolli nymphs and tadpole density were positive under both predator presence and absence. However, the slope under the presence of a predator was steeper than that under absence of the predator (“predator-by-tadpole density interaction” was significant). These results suggest that a higher tadpole density in June provides an abundant food resource for K. deyrolli nymphs and also moderates predation pressure from L. japonensis.     

Wing polymorphism in Japanese and foreign strains of the brown planthopper, Nilaparvata lugens

Great variations in nymphal density and wing-form relationships occur in field populations of the brown planthopper, Nilaparvata lugens (Stål) (Homoptera: Delphacidae). When tested under uniform laboratory conditions, half of the female populations collected from 20 locations in Japan exhibited higher ratios of macropters with increasing nymphal density, but one quarter were macropterous over broad ranges of density, both types being classified as group III. Three predominantly brachypterous populations (group I) were found, even under crowded conditions and one intermediate (group II).The populations from South Asian countries, i.e. Philippines (one), Indonesia (two) and Malaysia (one) belonged to group I, but the three populations from the coastal areas of China belonged to group II. Those from Taiwan differed in responses to density; one was of group II type, and two of group III type.From these comparisons, immigration routes of N. lugens to Japan were estimated, and survival strategy of this migratory species is discussed.

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Zusammenfassung In Feldpopulationen von Nilaparvata lugens (Stål) (Homoptera: Delphacidae) gibt es grosse Variationen in der Larvendichte und in den Flügelform-Beziehungen. Unter einheitlichen Laboratoriumsbedingungen wurden diese Verhältnisse geprüft. Von 20 japanischen Herkünften zeigte die Hälfte der Weibchenpopulationen mit zunehmender Larvendichte einen höheren Macropterenanteil; ein Viertel was über einen grossen Dichtebereich macropter; beide Typen werden als Gruppe III bezeichnet. Drei Populationen wurden gefunden, die auch bei hoher Dichte vorwiegend brachypter waren (Gruppe I). Einer Zwischengruppe (II) gehörte eine einzige Population an.Die Populationen aus südasiatischen Ländern (1 aus den Philippinen, 2 aus Indonesien un 1 aus Malaysia) gehörten zu Gruppe I, die 3 Populationen aus den Küstenregionen von China jedoch zu Gruppe II. Die Populationen aus Taiwan unterschieden sich in der Reaktion auf die Dichte: 1 gehörte zu Gruppe II and 2 zu Gruppe III.Von diesen Daten werden Einwanderungswege von N. lugens nach Japan abgeleitet und die Überlebensstrategie dieser Wanderart diskutiert.