Differences between wheat and rice in the enzymic properties of ribulose-1,5-bisphosphate carboxylase/oxygenase and the relationship to photosynthetic gas exchange

The kinetic parameters of ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase (EC 4.1.1.39) in wheat (Triticum aestivum L.) and rice (Oryza sativa L.) were determined by rapidly assaying the leaf extracts. The respective K _m and V _max values for carboxylase and oxygenase activities were significantly higher for wheat than for rice. In particular, the differences in the V _max values between the two species were greater. When the net activity of CO₂ exchange was calculated at the physiological CO₂-O₂ concentration from these kinetic parameters, it was 22% greater in wheat than in rice. This difference in the in-vitro RuBP-carboxylase/oxygenase activity between the two species reflected a difference in the CO₂-assimilation rate per unit of RuBP-carboxylase protein. However, there was no apparent difference in the CO₂-assimilation rate for a given leaf-nitrogen content between the two species. When the RuBP-carboxylase/oxygenase activity was estimated at the intercellular CO₂ pressure from the enzyme content and kinetic parameters, these estimated enzyme activities in wheat and rice were similar to each other for the same rate of CO₂ assimilation. These results indicate that the difference in the kinetic parameters of RuBP carboxylase between the two species was offset by the differences in RuBP-carboxylase content and conductance for a given leaf-nitrogen content.Abbreviations DTT dithiothreitol - EDTA ethylenediamine-tetraacetic - PAR photosynthetically active radiation - RuBP ribulose-1,5-bisphosphate     

Egg yolk lipoprotein,a new supplement for the growth of mammalian cells in serum-free medium

Egg yolk lipoprotein promoted growth of a wide variety of mammalian cell lines, including plasma-cytomas and epithelial cell lines, in serum-free medium. The lipoprotein was active for cell growth when used with insulin, transferrin, ethanolamine and selenite. The most active lipoprotein fraction (YLP-pI7.5) was purified to give a single peak by chromatofocusing and gel filtration, and was homogeneous on a 0.35% agarose gel electrophoretogram. The lipoprotein was characterised as a very low density lipoprotein with a protein content of only 1.3%. This lipoprotein had an optimal concentration of 300 g/ml (4 g protein/ml). It was easily separable from proteinous molecules secreted into the serum-free medium by the cells, since it floated on the surface of the medium after addition of ammonium sulfate, to precipitate protein, and centrifugation. An associated structure of lipid and protein seemed to be still necessary for the lipoprotein to exhibit a growth promoting activity.     

Asymmetric Reduction of 4-Oxoisophorone Using Immobilized Thermophilic Bacteria Under Conditions of Controlled Cell Growth

Asymmetric reduction of 2,6,6,-trimethyl-2-cyclohexene-l,4-dione (4-oxoisophrone) to (6R)-2,2,6-trimethyl-1,4-cyclohexane-dione((3R)-dihydro-4-oxoisophorone) was catalysed by immobilized thermophilic bacteria, Thermomonospora curvata JTS 321. Because of leakage of entrapped cells from gel beads during reactions using culture medium, we optimized the medium to allow the microbial conversion under conditions of controlled cell growth. Of the media screened, liver infusion medium was found to be the most suitable and microbial conversion was achieved without cell leakage from the immobilized gels. Immobilized T. curvata cells were repeatedly used for the asymmetric reduction of 4-oxoisophorone, more than 15 times, with an extent of conversion of 50%.     

Human hybrid immunoglobulin M containing immunoglobulin A

Summary Some hybridoma clones made by fusion of a human lymphoblastoid cell line, HO323 with human B lymphocytes, secreted not only IgA but also IgM-like immunoglobulin molecules. The IgM-like immunoglobulin had a molecular size of 900 K which corresponded to that of IgM. Immunochemical analyses revealed that the IgM-like immunoglobulin contained two monomeric IgA and three monomeric IgM molecules. In the IgA moieties, half of original light chains were replaced withx chains derived from the IgM, and vice versa.     

The atypical M2 segment of the beta subunit confers picrotoxinin resistance to inhibitory glycine receptor channels.

Purified preparations of the inhibitory glycine receptor (GlyR) contain alpha and beta subunits, which share homologous primary structures and a common transmembrane topology with other members of the ligand-gated ion channel superfamily. Here, a beta subunit-specific antiserum was shown to precipitate the [3H]strychnine binding sites localized on alpha subunits from membrane extracts of both rat spinal cord and mammalian cells co-transfected with alpha and beta cDNAs. Further, inhibition of alpha homo-oligomeric GlyRs by picrotoxinin, a non-competitive blocker of ion flow, was reduced 50- to 200-fold for alpha/beta hetero-oligomeric receptors generated by cotransfection. Site-directed mutagenesis identified residues within the second predicted transmembrane segment (M2) of the beta subunit as major determinants of picrotoxinin resistance. These data implicate the M2 segment in blocker binding to and lining of the GlyR chloride channel.     

Characterization of a glucuronyltransferase: neolactotetraosylceramide glucuronyltransferase from rat brain

The properties of a rat brain glucuronyltransferase, which is presumed to be associated with the biosynthesis of the HNK-1 epitope on sulfoglucuronyl glycolipids, are described. The enzyme required divalent cations for reaction, with maximal activity at 10mm Mn²⁺, and exhibited a dual optimum at pH 4–5 and pH 6 depending upon the buffer used, with the highest activity at pH 4.5 in MES buffer. This enzyme strictly recognized the Gal1-4GlcNAc terminal structure, and was highly specific for neolacto (type 2) glycolipids as acceptor. The enzyme was localized specifically in the brain, and was barely detected in other issues, including the thymus, spleen, liver, kidney, lung, and sciatic nerve fibres. Phosphatidylinositol and phosphatidylserine increased the enzymatic reaction 4.4- and 2.3-fold, respectively, whereas phosphatidylcholine slightly decreased the rate.Abbreviations GlcA glucuronic acid - Lc-PA₁₄ lactotetraose-phenyl-C₁₄H₂₉ - nLc-PA₁₄ neolactotetraose-phenyl-C₁₄H₂₉ - nLcOse₄-Cer neolactotetraosylceramide - NP-40 Nonidet P-40 - PC phosphatidylcholine - PE phosphatidylethanolamine - PI phosphatidylinositol - PS phosphatidylserine - SGGL sulfoglucuronyl glycolipid     

Enhancement of epidermal growth factor receptor expression on glioma cells by recombinant tumor necrosis factor α

Summary Recombinant tumor necrosis factor (rTNF; optimal dose 1000 U/ml) significantly increased the density of epidermal growth factor receptor (EGF-R) in three of four glioma cell lines in culture as determined by binding analysis of anti-EGF-R monoclonal antibody (mAb) 425. Since enhancement of EGF-R expression by rTNF- was inhibited when cells were treated with the protein synthesis inhibitor cycloheximide, the effects of rTNF may be protein-synthesis-dependent. The dose of rTNF that was optimal for up-regulation of EGF-R on glioma cells did not inhibit the growth of these cells.¹²⁵I-labeled mAb 425 lysed glioma cells in culture following its internalization into the cells. After glioma cells had been treated with rTNF, the growth-inhibitory effects of the mAb were significantly enhanced, probably a reflection of the increase in EGF-R density on the tumor cell surfaces. The rTNF effects were specific to the EGF-R and did not affect unrelated glioma-associated antigens. In our previous clinical trials,¹²⁵I-labeled mAb 425 showed immunotherapeutic effects in glioma patients. The present study provides the basis for considerations of combined immunotherapy of glioma patients with¹²⁵I-labeled mAb 425 and rTNF.     

Phototropic stimulation does not induce unequal distribution of indole-3-acetic acid in maize coleoptiles

Distribution of endogenous diffusible auxin into agar blocks from phototropically stimulated maize coleoptile tips was studied using a bioassay and a physicochemical assay, to clarify whether phototropism in maize coleoptiles involves a lateral gradient in the amount of auxin. At 50 min after the onset of phototropic stimulation, when the phototropic response was still developing, direct assay of the blocks with the Avena curvature test showed that the auxin activity in the blocks from the shaded half-tips was twice that of the lighted side, at both the first and second positive phototropic curvatures. However, physicochemical determination following purification showed that the amount of indole-3-acetic acid (IAA) was evenly distributed in the blocks from lighted and shaded coleoptile half-tips at both the first and second positive phototropic curvatures. The even distribution of the IAA was also confirmed with the Avena curvature test following purification by HPLC. These results indicate that phototropism in maize coleoptiles is not caused by a lateral gradient of IAA itself and thus cannot be described by the Cholodny-Went theory. Furthermore, the lower auxin activity in the blocks from the lighted half-tips suggests the presence of inhibitor(s) interfering with the action of auxin and their significant diffusion from unilaterally illuminated coleoptile tips.