Some Properties of Extracellular Acetylxylan Esterase Produced by the Yeast Rhodotorula mucilaginosa

The red yeast Rhodotorula mucilaginosa produced an esterase that accumulated in the culture supernatant on induction with triacetin. The enzyme was specific for substrates bearing an O-acetyl group, but was relatively nonspecific for the rest of the molecule, which could consist of a phenol, a monosaccharide, a polysaccharide, or an aliphatic alcohol. The esterase was more active against acetylxylan and glucose beta-d-pentaacetate than were a number of esterases from plant and animal sources, when activities on 4-nitrophenyl acetate were compared. The enzyme exhibited Michaelis-Menten kinetics and was active over a broad pH range (5.5 to 9.2), with an optimum between pH 8 and 10. In addition, the enzyme retained its activity for 2 h at 55 degrees C. The yeast that produced the enzyme did not produce xylanase and, hence, is of interest for the production of acetylxylan esterase that is free of xylanolytic activity.     

Analysis of Escherichia coli K12 F factor transfer genes: traQ, trbA, and trbB

The genes that encode the transfer properties of plasmid F, the fertility factor of Escherichia coli K12, are known to be clustered over a large, 33.3-kb segment of F DNA. As the central segment of the transfer region has not previously been well characterized, we constructed a detailed restriction map of the large F EcoRI DNA fragment, fl, and isolated a series of plasmid derivatives that carry various overlapping segments of this F tra operon DNA. We also analyzed the protein products of those clones that carried DNA segments extending over the region between traF and traH. This region was known to include traQ, a gene required for efficient conversion of the direct product of traA to the 7000-Da pilin polypeptide. We identified the traQ product as a polypeptide that migrates as a 12,500-Da protein on sodium dodecyl sulfate-polyacrylamide gels. We also detected the products of two other new genes that we have named trbA and trbB. These polypeptides migrate with apparent molecular weights of 14,200 and 18,400, respectively. Analysis of plasmid deletion derivatives that we constructed in vitro shows that these genes map in the order traF trbA traQ trbB traH. The presence of a plasmid carrying a small 0.43-kb fragment that expressed only the 12,500 traQ product caused the traA product of a co-resident compatible plasmid to be converted to the 7000-Da pilin polypeptide, demonstrating that TraQ is the only tra operon product required for this step of F-pilin biosynthesis.     

A model for glucose metabolism in energy-sufficient bacterial cultures

Summary A model for uncoupled glucose uptake under energy-sufficient conditions is presented. The model is derived from glucose catabolic pathways. The resulting model predicts specific glucose uptake rate as a function of both growth rate and extracellular glucose concentration. This prediction is consistent with reported literature data.     

Developmental changes of glucocorticoid receptors in the rat pancreas

Glucocorticoids are known to play a role in the maturation of the exocrine pancreas. The exact mechanism of glucocorticoid action in pancreatic ontogeny is, however, not clear. The present study characterized and quantitated the binding of [3H]dexamethasone to cytosol fractions from pancreata of rats at various ages. Trunk blood samples from these rats were also checked for levels of free and bound corticosterone. Specific and saturable bindings for dexamethasone were found in pancreatic cytosol fractions from newborn suckling and adult rats. Competition studies showed a preference for steroids with glucocorticoid activity. Specific binding was relatively low in pancreatic cytosol from newly born and 1-day old pups. A significant rise was seen after day 15. Cytosolic binding capacities were greatest from pancreata obtained from pups at weaning (3rd to 5th weeks). Values then declined toward the adult level. Scatchard analysis revealed a single class of binding sites with a dissociation constant (Kd) of 7.3 (+/- 1.1) X 10(-8) M and number of binding sites equalled to 1.29 (+/- 0.18) X 10(-13) mole/mg of cytosolic protein in adult rat pancreas. Pancreata from 25- and 15-day old rats had Kds of 3.4 (+/- 0.8) X 10(-8) M and 2.7 (+/- 0.7) X 10(-8) M with the number of binding sites equal to 1.77 (+/- 0.21) X 10(-13) mole/mg protein and 1.31 (+/- 0.16) X 10(-13) mole/mg protein respectively. Total plasma corticosterone concentration was low before day 10. It rose significantly by day 15, peaked at day 25, and then declined after weaning. About 5-15% of corticosterone during weaning and about 20-30% before and after weaning were in the free form. The peak level of dexamethasone binding corresponded to an increase in the plasma corticosterone level during weaning. This suggests a close relationship between plasma corticosterone levels and pancreatic glucocorticoid receptors. Both may, therefore, play a role in pancreatic development in the rat.     

Additive effects of apomorphine and clonidine on serotonin neurons in the dorsal raphe

Effects of apomorphine (APO) and clonidine (CLON) on the mesostriatal and mesolimbic serotonergic systems were examined in the present study. Both drugs selectively elevated serotonin (5-HT) concentrations in the dorsal raphe and the striatum without significantly altering 5-HT measures in the median raphe and the hippocampus. Apomorphine also increased tryptophan and 5-hydroxyindoleacetic acid (5-HIAA) levels in the dorsal raphe and 5-HIAA level in the striatum. Clonidine did not markedly alter tryptophan and 5-HIAA measures, while it decreased 5-HT turnover rate in both region, as indicated by the ratio of 5-HIAA/5-HT levels. Co-administration of APO and CLON, at doses of each drug exerted maximum effects on 5-HT alone, produced an additive effect on 5-HT in the dorsal raphe, while their effects on 5-HT and 5-HIAA in the striatum were counteracting each other. Effects of APO on 5-HT and 5-HIAA were attributed to the elevation of 5-HT precursor tryptophan, while effects of CLON on 5-HT and 5-HIAA were due to a decreased rate of 5-HT turnover. Therefore, the present results support the hypothesis that the additive effects of APO and CLON on dorsal raphe 5-HT are mediated through different receptors and neuropharmacological mechanisms.     

Effect of bacterial growth-inhibiting ingredients on the Ames mutagenicity of medicinal herbs

A solvent fractionation method was introduced to screen for mutagenicity in 10 medicinal herbs being consumed in Korea. The Ames mutagenicity test result of Scutellariae and Rhei was significantly increased by eliminating growth-inhibiting substances through solvent fractionation of the crude extract. It is suggested that a physicochemical pretreatment should reduce the false-negative results which are caused by the presence of growth-inhibiting substances in complex mixtures.     

Dietary induction of hepatic glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and malic enzyme in lean and obese female Zucker rats

Responses of the hepatic lipogenic enzymes, glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PGDH), and malic enzyme (ME) to starvation refeeding and diet shifting were determined in lean and obese female Zucker rats. Rats were either fed nonpurified diet, starved 48 hr, and then refed nonpurified diet or one of the refined carbohydrate diets containing either glucose, fructose, cornstarch, or sucrose for 72 hr, or shifted from nonpurified diet directly to one of the refined carbohydrate diets for 72 hr. Initial activities were greater in obese than lean rats for all three enzymes studied. Similar to other strains of female rats, lean Zucker rats failed to demonstrate a starve-refeed response when refed nonpurified diet. Obese female littermates showed a statistically significant increase in enzymes when refed a nonpurified diet. Both lean and obese female Zucker rats demonstrated increases in enzyme activities above controls when starved and refed any of the refined carbohydrate diets. The greatest responses were observed when female rats were starved and refed sucrose; activities increased 2.6- to 3.5-fold in lean and 3.0- to 4.3-fold in obese Zuckers. In lean females 50-70% of the starve-refeed response observed with G6PDH and ME can be accounted for by simply shifting from a nonpurified diet to the respective refined carbohydrate diet, whereas in obese females only 33-55% of the increase could be attributed to diet shifting. Plasma testosterone/estrogen ratios were consistently 1.5 times higher in obese than in lean female rats. This phenotypic difference may potentiate the heightened starve-refeed overshoot response observed in obese rats.     

Polyphosphate Formation from Pyrophosphate in Intact Cells of a Photosynthetic Bacterium, Rhodospirillum rubrum

The hypothesis that the photosynthetic bacterium, Rhodospirillumrubrum, may synthesize polyphosphate using pyrophosphate asa precursor has been tested. The cells were cultured in thepyrophosphate media, and the cell yield, cell shape, and thepolyphosphate content were determined and compared with thatgrown in the orthophosphate media. In the control experiment, the total amount of polyphosphateformed by the cultures grown in the pyrophosphate media wasfound to be up to 70% less than that grown in orthophosphatemedia. When the ATP formation was blocked by using the phosphorylationinhibitor, antimycin A, however, the cells grown in the pyrophosphatemedia formed up to 50% more polyphosphate than that grown inthe orthophosphate media. The results seem to support the hypothesis that Rhodospirillumrubrum may use pyrophosphate as the precursor of polyphosphate. (Received May 26, 1986; Accepted February 21, 1987)     

Electrical injury mechanisms: electrical breakdown of cell membranes

Electric fields are capable of damaging cells through both thermal and nonthermal mechanisms. While joule heating is generally recognized to mediate tissue injury in electrical trauma, the possible role of electrical breakdown of cell membranes has not been thoroughly considered. Evidence is presented suggestive that in many instances of electrical trauma the local electrical field is of sufficient magnitude to cause electrical breakdown of cell membranes and cell lysis. In theory, large cells such as muscle and nerve cells are more vulnerable to electrical breakdown. To illustrate the significance of cell size and orientation, a geometrically simple model of an elongated cell is analyzed.     

A freeze-fracture study of muscle fibres infected with Trichinella spiralis

The muscle fibres of mice containing the infective-stage larvae of the nematode Trichinella spiralis have been studied by means of the freeze-fracturing technique. The larva lies in what appears to be a fluid-filled cavity within the cytoplasm of an altered muscle fibre. There is no membrane separating the cytoplasm of the nurse cell from the cavity surrounding the larva which is therefore truly intracellular, unlike many parasites that reside within a membrane-lined parasitophorous vacuole within the host cell. This altered muscle fibre, known as a nurse cell, lacks myofilaments but does contain extensive cisternae of endoplasmic reticulum; membrane-bound vesicles are budded off from the endoplasmic reticulum and traverse the cytoplasm towards the cavity containing the nematode where they apparently pass into the cavity. It is suggested that the contents of these vesicles are used to sustain the nematode. Attention is drawn to the similarity to giant cells that have been induced by the plant-parasitic nematode Meloidogyne in the roots of host plants and which sustain the nematode. The conversion of the muscle fibre into a nurse cell is probably brought about by the presence of a metabolic sink, the larval nematode, within the cell. This take-over of the control of a metazoan cell by another metazoan organism is most unusual and warrants further study.