Conformational feature of neuroactive domoic acid: X-ray structural comparison with isodomoic acid A and alpha-kainic acid.

As an aid for developing a new type of potent insecticide acting on the neuromuscular junction, conformational characteristics of domoic acid and isodomoic acid A, the naturally occurring glutamate agonists, were investigated by X-ray crystal analyses. Conformational comparison with a neuroactive alpha-kainic acid provides information concerning the stereochemical feature responsible for the biological activity.     

The membrane-bound high-molecular-mass cytochromes c from Desulfovibrio gigas and Desulfovibrio vulgaris Hildenborough; EPR and Mössbauer studies

The high-molecular-mass cytochromes c (Hmcs) from the sulfate-reducing bacteria Desulfovibrio gigas and Desulfovibrio vulgaris (Hildenborough) were found to be strongly bound to the cytoplasmic membrane. After detergent solubilization they were shown to be water soluble and to be similar to those previously isolated from the soluble fractions in terms of N-terminal sequence, molecular mass, UV-visible and EPR spectroscopies. In D. gigas, higher amounts of Hmc can be obtained from the membranes than from the soluble fraction. This enabled further characterization of both cytochromes. The apparent heme reduction potentials of both Hmcs, determined at pH 7.5 through visible and EPR redox titrations, span a large range of redox potentials, approximately between 0 and –280?mV, and can be roughly divided into three groups: four to five hemes have E ⁰s of –30?mV to –100?mV, three to four hemes have E ⁰s around –170?mV, and seven to eight hemes have a lower E ⁰ of –250 to –280?mV. Several of these redox potentials are strongly pH dependent. Mössbauer studies of oxidized and reduced D. vulgaris Hmc show that this protein contains two high-spin hemes in both oxidation states. The rate of reduction of both Hmcs with the periplasmic hydrogenases from the corresponding organisms is extremely slow.     

Studies on siderophore production and effect of iron deprivation on the outer membrane proteins of Madurella mycetomatis

The purpose of this investigation was to determine whether Madurella mycetomatis, the most frequent agent of eumycotic mycetomas, produces siderophores and synthesizes new outer membrane proteins under iron-starvation conditions. Siderophore production, only of the hydroxamate type, was demonstrated in all nine strains tested. It was regulated by extracellular iron concentrations. Under iron-restricted conditions, M. mycetomatis expressed various outer membrane iron-regulated proteins, particularly of 24-kilodalton, that may participate in iron metabolism.     

A family with a satellited Yq chromosome

Summary A large pedigree with a satellited Yq chromosome is described, Q, C, and NOR banding were performed. Family C proband suffers from a Klinefelter syndrome.     

Calcium binding to phospholipid: structural study of calcium glycerophosphate.

To consider possible interaction of the phospholipid membrane with calcium ions, crystal structures of calcium dl-alpha- and beta-glycerophosphates (alpha- and beta-CaGs, respectively) were investigated by X-ray diffraction methods. After many attempts, relatively large single crystals of beta-CaG were prepared from the aqueous solution containing HCl, while crystals of CaHPO4.2H2O were obtained from alpha-CaG solution under the same crystallization conditions. The crystal structure of beta-CaG is orthorhombic with space group Pna2(1) and cell dimensions of a = 8.251(1), b = 13.038(3), c = 25.483 (10) A, V = 2741.5 (13) A3 and Z = 16 [four molecules (A to D) in an asymmetric unit]. Molecules of A to D took, as a whole, similar extended conformations, although A and B were different from C and D in the orientation about a glycerol C-C bond. Four independent beta-glycerophosphates commonly act as two types of bidentate ligands, where one is the coordination to the calcium ion by the glycerol O(1) and phosphate O(22) atoms, and the other by the phosphate O(22) and O(23) atoms, thus forming the calcium coordination of a distorted square plane, respectively. Each of four independent calcium ions forms the same coordination geometry of a distorted pentagonal bipyramid. Infinite double layers consisting of alternate A/B molecules and of alternative C/D ones and sandwiching calcium ions were arranged face-to-face along the b-direction and were piled up in the a-direction, thus forming the stacked bilayer unit with the thickness of d002 = 12.75 A. The elaborate networks of calcium coordinations and hydrogen bondings were formed among the layers and stabilized the crystal structure. Based on the structural parameters of the present beta-CaG crystal, a possible interaction model of phospholipid with calcium ions was proposed.     

X-ray crystallographic conformational study of 5'-O-[N-(L-alanyl)-sulfamoyl]adenosine, a substrate analogue for alanyl-tRNA synthetase

In order to elucidate the substrate specificity of alanyl-tRNA synthetase, 5'-O-[N-(L-alanyl)sulfamoyl]adenosine (Ala-SA), an analogue of alanyl-AMP, was chemically synthesized. Its binding ability is similar to that of the substrate based on the inhibitory activity for the aminoacylation of alanyl-tRNA synthetase. Taking advantage of the stable sulfamoyl bond of Ala-Sa, compared with the highly labile aminoacyl bond of alanyl-AMP, the molecular conformation of the former inhibitor was studied by X-ray single crystal analysis. Crystal data are as follows: C13H19N7O7S.2H2O, space group C2, a = 39.620(6), b = 5.757(1), c = 20.040(3) A, beta = 117.2(1) degrees, V = 4065(9) A3, Z = 8, and final R = 0.065 for 2785 independent reflections of F(2)0 greater than or equal to 2 sigma (F0)2. In the crystal, the molecule is in a zwitterionic state with the terminal amino group protonated and sulfamoyl group deprotonated, and takes an open conformation, where the L-alanine moiety is located far from the adenosine moiety with gauche/trans and trans orientations about the exocyclic C(4')-C(5') and C(5')-O(5') bonds, respectively. The conformation of the adenosine moiety is anti for the glycosyl bond and C(3')-endo for the ribose puckering, and alanine is in the usually observed trans region for the psi torsion angle. The molecular dimensions of the sulfamoyl group are nearly the same as those of the phosphate group. The biological significance of the observed Ala-SA conformation is discussed in relation with the molecular conformation of tyrosyl-AMP complexed with tyrosyl-tRNA synthetase.     

Dose-dependent hyporreactivity to phytohemagglutinin in systemic lupus erythematosus

The response of peripheral blood lymphocytes to stimulation with optimal and suboptimal doses of PHA was measured in patients with active SLE before initiation of therapy. The [³H]thymidine uptake of SLE patient's lymphocytes was significantly lower than that of their matched controls when cells were stimulated with suboptimal PHA doses in the presence of autologous plasma. A moderate improvement in the PHA response was observed by culturing washed patient's lymphocytes in medium supplemented with pooled normal human plasma, but only in one case the response reverted to normal values. A significant inhibitory effect of SLE plasma on the response of normal donor's lymphocytes to stimulation with low PHA doses, which was independent from the presence of lymphocytotoxic antibodies and persisted after complement inactivation was observed in further experiments.The results indicate that depression of lymphocyte transformation could be demonstrated in patients with active SLE using suboptimal doses of PHA and suggest that this depression may be caused by both a defect in the responding lymphocyte populalation and the presence of inhibitory factor(s) in SLE plasma.     

Production of nattokinase by high cell density fed-batch culture of Bacillus subtilis

Bacillus subtilis was cultivated to high cell density for nattokinase production by pH-stat fed-batch culture. A concentrated mixture solution of glucose and peptone was automatically added by acid-supplying pump when culture pH rose above high limit. Effect of the ratio of glucose to peptone in feeding solution was investigated on cell growth and nattokinase production by changing the ratio from 0.2 to 5 g glucose/g peptone. The highest cell concentration was 77 g/L when the ratio was 0.2 g glucose/g peptone. Cell concentration decreased with increasing the ratio of glucose to peptone in feeding solution, while the optimum condition existed for nattokinase production. The highest nattokinase activity was 14,500 unit/mL at a ratio of 0.33 g glucose/g peptone, which was 4.3 times higher than that in batch culture.