Continuous production of L-tryptophan from indole and L-serine by immobilized Escherichia coli cells

Escherichia coli B 10, which has high activity of tryptophan synthetase, was grown in a 50-L batch culture in order to determine in which growth phase the cells have the highest specific tryptophan productivity. Accordingly, whole cells of the stationary phase were used for immobilization in polyacrylamide beads. After immobilization, these immobilized cells had 56% activity of tryptophan synthetase compared with that of free cells. First, the properties of immobilized cells were investigated. Next, discontinuous productions of L-tryptophan were carried out by using immobilized cells. In discontinuous production of L-tryptophan by the batch, the activity remaining of immobilized cells was 76-79% after 30 times batchwise use. In continuous production of L-tryptophan with a continuous stirred tank reactor (CSTR), the activity remaining of the immobilized cells was 80% after continuous use for 50 days. The maximum productivity of L-tryptophan in this CSTR system was 0.12 g tryptophan L(-1) h(-1).     

Evidence for an oscillator other than luteinizing hormone controlling the secretion of progesterone in cattle

The objective of this study was to quantify and compare the frequencies of pulses in ovarian and systemic concentrations of progesterone, systemic concentrations of luteinizing hormone (LH) and rate of ovarian blood flow. Blood was collected simultaneously from previously implanted catheters in the ovarian venous circulation and jugular vein on Day 12 or 13 of estrous cycles from 4 nonlactating dairy cows. Blood was collected at a rate of 2.5 ml/min for 5 min out of every 10 min over an 8-h period. The mean rate of blood flow in the ovarian artery during the 5-min collection period was estimated by an electromagnetic blood flow transducer. Pulses were observed over time in both ovarian and systemic concentrations of progesterone at frequencies that ranged between 0.625 and 0.875 cycles/h (1.1 to 1.5 h/cycle) among the animals. Only one or two episodes of release of LH were observed during the 8-h period, and transient increases in blood flow to the ovaries were associated temporally with each episode of LH release. The estimated frequencies for release of LH and increased blood flow were the same for each animal and ranged between 0.250 and 0.375 cycles/h. A second cycle with a frequency similar to that for LH was evident in the spectral density functions for ovarian and systemic concentrations of progesterone. This cycle was eliminated when the cycle for LH was removed from the data for progesterone, but the magnitude and frequency of the pulses in progesterone were not affected.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reduced synthesis of basement membrane heparan sulfate proteoglycan in streptozotocin-induced diabetic mice

In diabetes, certain basement membranes become thicker yet more porous than normal. To identify possible changes in the basement membrane, we have grown the Engelbreth-Holm-Swarm tumor, a tissue that produces quantities of basement membrane in normal mice and in streptozotocin-treated, insulin-deficient, diabetic mice. The level of laminin, a basement membrane-specific glycoprotein, and the level of total protein were slightly elevated in the diabetic tissue. In contrast, the level of the basement membrane specific heparan sulfate proteoglycan was only 20% of control. The synthesis of this proteoglycan was also reduced in the diabetic animals, while the synthesis of other proteoglycans by tissues such as cartilage was normal. The synthesis of the heparan sulfate proteoglycan in diabetic animals was inversely related to plasma glucose levels showing an abrupt decrease above the normal range of plasma glucose. Insulin restored synthesis to normal but this required doses of insulin that maintained plasma glucose at normal levels for several hours. Since the heparan sulfate proteoglycan in the basement membrane restricts passage of proteins, its absence could account for the increased porosity of basement membrane in diabetes. A compensatory synthesis of other components could lead to their increased deposition and the accumulation of basement membrane in diabetes.     

In vitro and in vivo regulation of chloroplast glyceraldehydes-3-phosphate dehydrogenase isozymes from Chenopodium rubrum. I. Purification and properties of isozymes

Chloroplast glyceraldehyde-3-phosphate dehydrogenase (GPD, EC 1.2.1.13) was purified from leaves of Chenopodium rubrum L. Aggregated (≥ 10⁶) and disaggregated (165 × 10³) molecular weight forms were obtained by gel filtration in the presence of NAD⁺ and NADP⁺, respectively. The disaggregated enzyme was separated into two isozymes by inverse ammonium sulphate gradient solubilization: "NADP-GPD I" was homotetrameric (subunit molecular weight 39 × 10³); "NADP-GPD II" was heterotetrameric (subunit molecular weights 39 × 10³ and 43 × 10³). Isoelectric focusing of the isozymes, both aggregated and disaggregated, revealed two isoelectric forms in each case, at 4.3 and 7.7. Chloroplast GPD was "NADP-suppressed" in crude extracts due to partial oxidation, incubation with dithioerythritol restored full activity.     

Direct effects on the membrane potential due to "pumps" that transfer no net charge

The effects of active ionic transport are included in the derivation of a general expression for the zero current membrane potential. It is demonstrated that an active transport system that transfers no net charge (nonrheogenic) may, nevertheless, directly alter the membrane potential. This effect depends upon the exchange of matter within the membrane between the active and passive diffusion regimes. Furthermore, in the presence of such exchange, the transmembrane active fluxes measured by the usual techniques and the local pumped fluxes are not identical. Several common uses of the term “electrogenic pump” are thus shown to be inconsistent with each other. These inconsistencies persist when the derivation is extended to produce a Goldman equation modified to account for active transport; however, that equation is shown to be limited by less narrow constraints on membrane heterogeneity and internal electric field than those previously required. In particular, it is applicable to idealized mosaic membranes limited by these requirements.     

Lipid Composition of Purified Vesicular Stomatitis Viruses

Methods are described for the production of vesicular stomatitis (VS) virus of sufficient purity for reliable chemical analysis. VS virions released from infected cells were concentrated and purified at least 150-fold by sequential steps of precipitation with polyethylene glycol, column chromatography, rate zonal centrifugation, and equilibrium centrifugation. The Indiana serotype (VS(Ind) virus) propagated in L-cells was found to contain 3% ribonucleic acid, 64% protein, 13% carbohydrate, and 20% lipid; the molar ratio of cholesterol to phospholipid was 0.6 or greater. Thin-layer chromatography revealed no unusual neutral lipids or phospholipids and gas-liquid chromatography revealed no unusual fatty acids incorporated into VS virions. The antigenically distinct New Jersey serotype (VS(NJ) virus) grown in L-cells showed a similar lipid profile except that the proportion of neutral lipids was larger than in VS(Ind) virus also grown in L-cells. This differences was less pronounced when the lipid composition of VS(Ind) and VS(NJ) viruses grown in chick embryo cells was compared, but VS(NJ) virus grown in either cell type always contained larger amounts of neutral lipids other than cholesterol than did VS(Ind) virus. The lipid composition of both VS(Ind) and VS(NJ) viruses grown in L-cells or chick embryo cells more closely resembled that of plasma membrane than of whole cells. A consistent finding was the relatively large amounts of phosphatidylethanolamine and sphingomyelin and the relatively small amounts of phosphatidylcholine in both VS viruses compared with uninfected whole L-cells and chick embryo cells or their plasma membranes. The methods available for isolation of plasma membranes were inadequate for conclusive comparison of the lipids of VS virions with the lipids of the plasma membranes of their host cells. Nevertheless, the data obtained are consistent with two hypotheses: (i) the lipid composition of VS viruses primarily reflects their membrane site of maturation, and (ii) the newly synthesized viral proteins inserted into cell membranes influence the proportions of phospholipids and neutral lipids selected for incorporation into the viral membrane.