Physiological adaptations of anaerobic bacteria to low pH: metabolic control of proton motive force in Sarcina ventriculi.

Detailed physiological studies were done to compare the influence of environmental pH and fermentation end product formation on metabolism, growth, and proton motive force in Sarcina ventriculi. The kinetics of end product formation during glucose fermentation in unbuffered batch cultures shifted from hydrogen-acetate production to ethanol production as the medium pH dropped from 7.0 to 3.3. At a constant pH of 3.0, the production of acetate ceased when the accumulation of acetate in the medium reached 40 mmol/liter. At a constant pH of 7.0, acetate production continued throughout the entire growth time course. The in vivo hydrogenase activity was much higher in cells grown at pH 7.0 than at pH 3.0. The magnitude of the proton motive force increased in relation to a decrease of the medium pH from 7.5 to 3.0. When the organism was grown at pH 3.0, the cytoplasmic pH was 4.25 and the organism was unable to exclude acetic acid or butyric acid from the cytoplasm. Addition of acetic acid, but not hydrogen or ethanol, inhibited growth and resulted in proton motive force dissipation and the accumulation of acetic acid in the cytoplasm. The results indicate that S. ventriculi is an acidophile that can continue to produce ethanol at low cytoplasmic pH values. Both the ability to shift to ethanol production and the ability to continue to ferment glucose while cytoplasmic pH values are low adapt S. ventriculi for growth at low pH.     

Molecular cloning of a cDNA for the E1 alpha subunit of rat liver branched chain alpha-ketoacid dehydrogenase

We have isolated a cDNA encoding the branched chain alpha-ketoacid dehydrogenase E1 alpha subunit. A rat liver lambda gt11 expression library was screened with antibody reactive with the 2-oxoisovalerate dehydrogenase (lipoamide) component. A positive clone, lambda BZ304, contains a 1.7-kilobase pair cDNA insert with a 1323-base pair open reading frame. Translation of the open reading frame predicts the 24 residues of the previously reported phosphorylation sites 1 and 2 for the bovine kidney and rabbit heart enzymes. The N-terminal sequence of purified E1 alpha was determined, and this sequence was found 40 residues from the beginning of the deduced peptide sequence. Northern blots of rat liver and muscle RNA demonstrate a single mRNA species of approximately 1.8 kilobase pairs in each tissue, suggesting that this cDNA is nearly full length.     

The purification of an erythroid protein which binds to enhancer and promoter elements of haemoglobin genes.

An erythroid nuclear protein (EF1), originally detected as a protein binding within the nuclease hypersensitive site upstream of the chicken beta H-globin gene, has been purified. This protein of 37,000-39,000 molecular weight binds to three sites within the hypersensitive region: one between the CCAAT and TATA boxes, the second (further upstream) next to a NF1 binding site, and the third adjacent to a regulatory element found in a number of beta-globin genes. The EF1 protein also binds to an erythroid-specific promoter element of the mouse alpha-globin gene and to two sites within the chicken beta A-globin enhancer. These six EF1-binding sites are related by the consensus sequence A/TGATAA/GG/C. A minor protein of molecular weight 72,000 which co-purifies with EF1 also binds to the same sequences.     

Effects of calcium input/output on the stability of a system for calcium-regulated viscoelastic strain fields

The Goodwin and Trainor model of pattern generation in calcium-regulated strain fields is studied in the case where calcium input and calcium output processes are involved. It is shown that the properties of the original model may still remain provided that the input-output processes are not unstable. In this last case, despite the eventual stabilizing effect of the calcium exchange term, perturbations of the generalized system can grow and lead to inhomogeneous solutions. Applications to cell differentiation and cell growth are discussed.     

Sugar uptake by maize endosperm suspension cultures

Maize (Zea mays L.) endosperm suspension cultures are a useful model system for studying biochemical and physiological events in developing maize endosperm. In this report, sugar uptake by the cultures is characterized. Uptake of ¹⁴C-labeled fructose and l-glucose was linear with time, while the rate of uptake of radioactivity from sucrose increased over a 120 min period. Both saturable and linear components of uptake were observed for fructose, glucose, sucrose, 1′-deoxy-1′-fluorosucrose, and maltose. Uptake of mannitol, sorbitol, and l-glucose took place at lower rates and was linear with concentration. Rates of incorporation of radioactivity from fructose and glucose exceeded that of sucrose at all concentrations tested. Kinetics of 1′-deoxy-1′-fluorosucrose uptake indicated that ¹⁴C from sucrose can be taken up by a saturable carrier of intact sucrose as well as by invertase hydrolysis and subsequent uptake of hexoses. Cell wall invertase was demonstrated histochemically. Further study of fructose uptake at a concentration at which the saturable component predominated revealed sensitivity to metabolic inhibitors, respiratory uncouplers, the nonpermeant sulfhydryl reagent p-chloromercuribenzenesulfonic acid, and nigericin. Uptake was not affected by valinomycin plus K⁺ and was stimulated by fusicoccin. Fructose and glucose uptake was not pH-sensitive below pH 7.0, whereas uptake of radioactivity from sucrose and 1′-deoxy-1′-fluorosucrose declined as the pH was increased above 5.0. Fructose uptake was not completely inhibited by glucose and vice versa, suggesting the presence of specific carriers. These results indicate that maize endosperm suspension cultures (a) absorb fructose via a typical, energy-requiring, carrier-mediated proton cotransport system; (b) possess saturable carriers for glucose and sucrose; and (c) also absorb sucrose via hexose uptake after sucrose hydrolysis by extracellular invertase.     

Monoclonal antibody-based immunoassay of vitellogenin in the blood of male channel catfish (Ictalurus punctatus).

1. A monoclonal antibody to vitellogenin of channel catfish (Ictalurus punctatus) was made, and its specificity was demonstrated using Western blots of serum from female fish, estradiol-treated male fish, untreated male fish, vitellogenin purified by three different methods and egg extracts. 2. An enzyme-linked immunosorbent assay (ELISA), using this monoclonal antibody, detected vitellogenin in the plasma of 59 out of 60 untreated 17-24-month-old male channel catfish with a mean concentration of 338 micrograms/ml and a maximum concentration of 4240 micrograms/ml. 3. Vitellogenin levels in male channel catfish were unrelated to testicular stage, gonadosomatic index and month.     

Subcutaneous narcotic infusions for cancer pain: treatment outcome and guidelines for use.

OBJECTIVE: To provide guidelines for the institution and maintenance of a continuous subcutaneous narcotic infusion program for cancer patients with chronic pain through an analysis of the narcotic requirements and treatment outcomes of patients who underwent such therapy and a comparison of the costs of two commonly used infusion systems. DESIGN: Retrospective study. SETTING: Tertiary care facilities and patients'' homes. PATIENTS: Of 481 patients seen in consultation for cancer pain between July 1987 and April 1990, 60 (12%) met the eligibility criteria (i.e., standard medical management had failed, and they had adequate supervision at home). INTERVENTION: Continuous subcutaneous infusion with hydromorphone hydrochloride or morphine started on an inpatient basis and continued at home whenever possible. OUTCOME MEASURES: Patient selectivity, narcotic dosing requirements, discharge rate, patient preference for analgesic regimen, side effects, complications and cost-effectiveness. RESULTS: The mean initial maintenance infusion dose after dose titration was almost three times higher than the dose required before infusion (hydromorphone or equivalent 6.2 v. 2.1 mg/h). Eighteen patients died, and the remaining 42 were discharged home for a mean of 94.4 (standard deviation 128.3) days (extremes 12 and 741 days). The mean maximum infusion rate was 24.1 mg/h (extremes 0.5 and 180 mg/h). All but one of the patients preferred the infusion system to their previous oral analgesic regimen. Despite major dose escalations nausea and vomiting were well controlled in all cases. Twelve patients (20%) experienced serious systemic toxic effects or complications; six became encephalopathic, which necessitated dose reduction, five had a subcutaneous infection necessitating antibiotic treatment, and one had respiratory depression. The programmable computerized infusion pump was found to be more cost-effective than the disposable infusion device after a break-even point of 8 months. CONCLUSIONS: Continuous subcutaneous infusion of opioid drugs with the use of a portable programmable pump is safe and effective in selected patients who have failed to respond to standard medical treatment of their cancer pain. Dose titration may require rapid dose escalation, but this is usually well tolerated. For most communities embarking on such a program a programmable infusion system will be more cost-effective than a disposable system.