Effects of chemical modification of carboxyl groups in the hemolytic lectin CEL-III on its hemolytic and carbohydrate-binding activities

Effects of chemical modification of carboxyl groups in the hemolytic lectin CEL-III on its activities were investigated. When carboxyl groups were modified with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and glycine methyl ester, hemolytic activity of CEL-III decreased as the EDC concentration increased, accompanied by reduction of oligomerization ability and hemagglutinating activity. However, binding ability of CEL-III for immobilized lactose was retained fairly well after modification, suggesting that one of two carbohydrate-binding sites might be responsible for such inactivation of CEL-III.     

Synthesis of the novel analogues of dysidiolide and their structure-activity relationship

The novel analogues of natural cdc25A inhibitor dysidiolide were synthesized. To investigate the structure-activity relationship, the inhibitory activity to enzyme and cell cycle was examined.     

Protection against malaria by anti-erythropoietin antibody due to suppression of erythropoiesis in the liver and at other sites

We have previously reported that erythropoiesis commences in the liver and spleen after malarial infection, and that newly generated erythrocytes in the liver are essential for infection of malarial parasites as well as continuation of infection. At this time, erythropoietin (EPO) is elevated in the serum. In the present study, we administered EPO or anti-EPO antibody into C57BL/6 (B6) mice to modulate the serum level of EPO. When mice were infected with a non-lethal strain (17NXL) of Plasmodium yoelii (blood-stage infection of 10(4) parasitized erythrocytes per mouse), parasitemia continued for 1 month, showing a peak at day 17. Daily injection of EPO (200 IU/day per mouse) from day five to day 14 prolonged parasitemia, whereas injection of anti-EPO antibody (1.5 mg/day per mouse) every second day from day five to day 28 decreased it. Erythropoiesis was confirmed in the liver, spleen and bone marrow by the appearance of nucleated erythrocytes (TER119+). When anti-EPO antibody was injected by the same protocol into mice infected with a lethal strain (17XL) of P. yoelii, all mice showed decreased parasitemia and recovered from the infection. These results suggest that the use of anti-EPO antibody after malarial infection may be of therapeutic value in severe cases of malaria.     

Role of L-ficolin/mannose-binding lectin-associated serine protease complexes in the opsonophagocytosis of type III group B streptococci

Serotype III group B streptococci (GBS) are a common cause of neonatal sepsis and meningitis. Although deficiency in maternal capsular polysaccharide (CPS)-specific IgG correlates with susceptibility of neonates to the GBS infection, serum deficient in CPS-specific IgG mediates significant opsonophagocytosis. This IgG-independent opsonophagocytosis requires activation of the complement pathway, a process requiring the presence of both Ca(2+) and Mg(2+), and is significantly reduced by chelating Ca(2+) with EGTA. In these studies, we defined a role of L-ficolin/mannose-binding lectin-associated serine protease (MASP) complexes in Ca(2+)-dependent, Ab-independent opsonophagocytosis of serotype III GBS. Incubation of GBS with affinity-purified L-ficolin/MASP complexes and C1q-depleted serum deficient in CPS-specific Ab supported opsonophagocytic killing, and this killing was inhibited by fluid-phase N-acetylglucosamine, the ligand for L-ficolin. Binding of L-ficolin was proportional to the CPS content of individual strains, and opsonophagocytic killing and C4 activation were inhibited by fluid-phase CPS, suggesting that L-ficolin binds to CPS. Sialic acid is known to inhibit alternative complement pathway activation, and, as expected, the bactericidal index (percentage of bacteria killed) for individual strains was inversely proportional to the sialic acid content of the CPS, and L-ficolin-initiated opsonophagocytic killing was significantly increased by addition of CPS-specific IgG2, which increased activation of the alternative pathway. We conclude that binding of L-ficolin/MASP complexes to the CPS generates C3 convertase C4b2a, which deposits C3b on GBS. C3b deposited by this lectin pathway forms alternative pathway C3 convertase C3bBb whose activity is enhanced by CPS-specific IgG2, leading to increased opsonophagocytic killing by further deposition of C3b on the GBS.     

Apoptosis induction by lectin isolated from the mushroom Boletopsis leucomelas in U937 cells

A 15-kDa lectin was isolated from the edible mushroom Kurokawa by affinity chromatography using N,N'-diacetylchitobiose-Sepharose 4B. The results of microsequencing analysis indicated that the lectin has a partial amino acid sequence similar to the mushroom lectin, Agaricus bisporus agglutinin (ABA). We found that the Kurokawa lectin inhibited proliferation of human monoblastic leukemia U937 cells dose-dependently. Several lines of evidence indicated that this inhibition was due to its apoptosis induction. We observed that the lectin induced apoptotic bodies formation, chromatin condensation, and DNA ladder formation, features of apoptosis. The DNA ladder formation was inhibited by a general inhibitor of caspases, which are known to play essential roles in apoptosis. In contrast, ABA did not have cell growth-inhibiting or apoptosis-inducing activities. Thus, the Kurokawa lectin is the first mushroom lectin with apoptosis-inducing activity.     

Pituitary adenylate cyclase-activating polypeptide enhances Ca(2+)-dependent neurotransmitter release from PC12 cells and cultured cerebellar granule cells without affecting intracellular Ca(2+) mobilization

Peroxisome proliferator-activated receptor gamma (PPAR gamma) belongs to a nuclear receptor super family that functions as a master regulator of adipocyte differentiation. PPAR gamma binds its DNA response element together with a partner, retinoid X receptor (RXR), in fat cells. Five RXR ligands (HX600, HX630, DA022, DA124, LGD1069, referred to as retinoid synergists) by themselves exhibit weak transactivation activity on the PPAR gamma response element. However, addition of PPAR gamma-specific ligand in this assay gave rise to a 5- to 13-fold increase, indicating a strong synergy between these ligands. LGD1069 was the most effective activator of the RXR/PPAR gamma heterodimer on the transactivation of the reporter gene. But, in contrast to the other four RXR ligands, LGD1069 did not show synergistic induction of ST 13 preadipocytes to adipocytes. This apparent contradiction may result from the ligand-binding property of LGD1069. In this article we discuss the fact that retinoid synergists also act as PPAR gamma synergists.     

Hepatic fatty acid-binding proteins of a teleost, Lateolabrax japonicus. The primary structures and location of a disulfide bond

Two fatty acid-binding proteins (FABP), FABP-1 and FABP-2, were purified from the liver cytosol of the teleost, Lateolabrax japonicus (Japan sea bass), and characterized. The complete primary structure of FABP-2 was determined by protein analysis to be the following: MDFSGTWQVY AQENYEEFLR AMELPADVIK MAKDIKPITE IKQSGNDFVV TSKTPGKTVT NSFTIGKEAD ITTMDGKKIR CVVNLEGGKL VCNTGKFCHI QELRGGEMVE TLTMGSTTLI RKSKKM. Partial peptide sequences of FABP-1 were also determined. Phylogenetic analysis indicates that FABP-2 is a homologue of mammalian hepatic FABP, whereas FABP-1 is most similar to the members of mammalian cardiac FABP subfamily. L. japonicus FABP-2 contains three cysteine residues, and a disulfide bond is identified between Cys-81 and Cys-92. A theoretical model of FABP-2 generated by a homology modeling method indicates close proximity of the two cysteine residues in the three-dimensional structure. This is a rather rare case of cytosolic protein having a disulfide bond under the normally reducing conditions of the cytosol, though the presence or absence of disulfide bonds does not seem to affect the ligand-binding ability.