Enzymic saccharification of sugar beet pulp

Culture filtrates of Talaromyces emersonii UCG 208 grown on beet pulp can convert the polysaccharide components of this agricultural waste to soluble sugars. The saccharification process is facilitated if the pulp is milled or incubated with alkali or peracetic acid before addition of enzyme. However, treatment of unmilled pulp with commercial pectinase prior to incubation with Talaromyces filtrate is also very effective; under suitable conditions, complete hydrolysis of total polysaccharides has been achieved.     

Production and Partial Purification of Antibiotic Materials Formed by Physarum gyrosum

Pure cultures of Physarum gyrosum were grown on agar plates with autoclaved Escherichia coli suspensions as the growth medium. Portions of such agar, after growth of the slime mold, contained diffusible materials that inhibited the growth of Bacillus subtilis, B. cereus, E. coli, Staphylococcus aureus, and Pseudomonas aeruginosa. Paper chromatography of extracts of such cultures revealed at least three different active fractions. Preliminary fractionations increased the specific activity by one order of magnitude, probably in part by removal of inactive material and in part by separating active components. The fractionations also demonstrated the multiplicity of the antibiotic activity. Fractions variously obtained always retarded the growth of the bacterial species listed above.     

Isolation of mutants of Talaromyces emersonii CBS 814.70 with enhanced cellulase activity

By a combination of genetic mutation and modification of growth medium the cellulase [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4 etc.] activity of culture filtrates of Talaromyces emersonii CBS 814.70 has been increased four-fold to approximately 2 U ml^?1 and a productivity of 20–25 Ul^?1h^?1. At 50°C this system was completely stable for at least 24 h. At 60°C in static reaction mixtures 19% of the original activity was lost compared with 21% when mixtures were shaken. At 70°C the loss of activity after 4 h was 64% without shaking and 70% when shaken. The cellulase system from Trichoderma reesei was decidedly less stable than that of Talaromyces emersonii under each of the above conditions. The ability of each enzyme system, separately and together, to digest beet pulp was investigated.     

SIRT1 Activation by Small Molecules: KINETIC AND BIOPHYSICAL EVIDENCE FOR DIRECT INTERACTION OF ENZYME AND ACTIVATOR?

SIRT1 is a protein deacetylase that has emerged as a therapeutic target for the development of activators to treat diseases of aging. SIRT1-activating compounds (STACs) have been developed that produce biological effects consistent with direct SIRT1 activation. At the molecular level, the mechanism by which STACs activate SIRT1 remains elusive. In the studies reported herein, the mechanism of SIRT1 activation is examined using representative compounds chosen from a collection of STACs. These studies reveal that activation of SIRT1 by STACs is strongly dependent on structural features of the peptide substrate. Significantly, and in contrast to studies reporting that peptides must bear a fluorophore for their deacetylation to be accelerated, we find that some STACs can accelerate the SIRT1-catalyzed deacetylation of specific unlabeled peptides composed only of natural amino acids. These results, together with others of this study, are at odds with a recent claim that complex formation between STACs and fluorophore-labeled peptides plays a role in the activation of SIRT1 (Pacholec, M., Chrunyk, B., Cunningham, D., Flynn, D., Griffith, D., Griffor, M., Loulakis, P., Pabst, B., Qiu, X., Stockman, B., Thanabal, V., Varghese, A., Ward, J., Withka, J., and Ahn, K. (2010) J. Biol. Chem. 285, 8340–8351). Rather, the data suggest that STACs interact directly with SIRT1 and activate SIRT1-catalyzed deacetylation through an allosteric mechanism.     

Identification and synthesis of major metabolites of Vasopressin V2-receptor agonist WAY-151932, and antagonist, Lixivaptan

Small molecule agonists and antagonists of the V(2)-vasopressin receptor have been discovered and have undergone clinical trials. In conjunction with these discovery programs, the synthesis and biological testing of various metabolites associated with these clinical targets were actively pursued. We now report the results of our synthetic efforts and the corresponding biological data generated for several of the metabolites of WAY-151932 and CL-347985 (Lixivaptan).     

Spatio-temporal relief from hypoxia and production of reactive oxygen species during bud burst in grapevine (Vitis vinifera)

Background and Aims Plants regulate cellular oxygen partial pressures (pO₂), together with reduction/oxidation (redox) state in order to manage rapid developmental transitions such as bud burst after a period of quiescence. However, our understanding of pO₂ regulation in complex meristematic organs such as buds is incomplete and, in particular, lacks spatial resolution.Methods The gradients in pO₂ from the outer scales to the primary meristem complex were measured in grapevine (Vitis vinifera) buds, together with respiratory CO₂ production rates and the accumulation of superoxide and hydrogen peroxide, from ecodormancy through the first 72 h preceding bud burst, triggered by the transition from low to ambient temperatures.Key Results Steep internal pO₂ gradients were measured in dormant buds with values as low as 2·5 kPa found in the core of the bud prior to bud burst. Respiratory CO₂ production rates increased soon after the transition from low to ambient temperatures and the bud tissues gradually became oxygenated in a patterned process. Within 3 h of the transition to ambient temperatures, superoxide accumulation was observed in the cambial meristem, co-localizing with lignified cellulose associated with pro-vascular tissues. Thereafter, superoxide accumulated in other areas subtending the apical meristem complex, in the absence of significant hydrogen peroxide accumulation, except in the cambial meristem. By 72 h, the internal pO₂ gradient showed a biphasic profile, where the minimum pO₂ was external to the core of the bud complex.Conclusions Spatial and temporal control of the tissue oxygen environment occurs within quiescent buds, and the transition from quiescence to bud burst is accompanied by a regulated relaxation of the hypoxic state and accumulation of reactive oxygen species within the developing cambium and vascular tissues of the heterotrophic grapevine buds.     

The abundant NK cells in human secondary lymphoid tissues require activation to express killer cell Ig-like receptors and become cytolytic

Natural killer cells are important cytolytic cells in innate immunity. We have characterized human NK cells of spleen, lymph nodes, and tonsils. More than 95% of peripheral blood and 85% of spleen NK cells are CD56(dim)CD16(+) and express perforin, the natural cytotoxicity receptors (NCRs) NKp30 and NKp46, as well as in part killer cell Ig-like receptors (KIRs). In contrast, NK cells in lymph nodes have mainly a CD56(bright)CD16(-) phenotype and lack perforin. In addition, they lack KIRs and all NCR expression, except low levels of NKp46. The NK cells of tonsils also lack perforin, KIRs, NKp30, and CD16, but partially express NKp44 and NKp46. Upon IL-2 stimulation, however, lymph node and tonsilar NK cells up-regulate NCRs, express perforin, and acquire cytolytic activity for NK-sensitive target cells. In addition, they express CD16 and KIRs upon IL-2 activation, and therefore display a phenotype similar to peripheral blood NK cells. We hypothesize that IL-2 can mobilize the NK cells of secondary lymphoid tissues to mediate natural killing during immune responses. Because lymph nodes harbor 40% and peripheral blood only 2% of all lymphocytes in humans, this newly characterized perforin(-) NK cell compartment in lymph nodes and related tissues probably outnumbers perforin(+) NK cells. These results also suggest secondary lymphoid organs as a possible site of NK cell differentiation and self-tolerance acquisition.