An assay for the measurement of the protein content of cells immobilized in carrageenan

1984 ◽  
Vol 30 (4) ◽  
pp. 475-481 ◽  
Author(s):  
A. Jones ◽  
T. Razniewska ◽  
B. H. Lesser ◽  
R. Siqueira ◽  
D. Berk ◽  
...  
Keyword(s):  
Protein Content ◽  
Immobilized Cells ◽  
Fine Powder ◽  
Cellular Protein ◽  
Cell Protein ◽  
Specific Productivity ◽  
Immobilized Cell ◽  
Freeze Dried ◽  
Lowry Method ◽  
Reproducible Method

A reliable and reproducible method for the estimation of the protein content of fungal cells immobilized in a carrageenan gel is described. The procedure depends upon the acid lability of the polysaccharide gel at 90 °C and on the acetone solubility of accumulated phenolics. Freeze-dried gel beads (2–3 mm) containing entrapped cells of Penicillium urticae were ground to a fine powder and samples of powder (~20 mg) were sequentially extracted with hot 1 N HCl – 0.9% NaCl and acetone. The precipitated residue contained the cell protein, which was then solubilized with 1 N NaOH at 90 °C and quantitated by the Folin–Lowry method. Interferences from both carrageenan and phenols were thus eliminated. The presence of carrageenan (20–25 mg) did not affect the recovery of varying amounts (0–2500 μg) of bovine serum albumin. The recovery of radiolabeled protein from immobilized cells was parallel to that of Folin–Lowry positive material over a range of 0–60 beads (0–60 mg powder). Cycloheximide (0–100 μg/mL) was shown to progressively inhibit the incorporation of L-[U-14C]leucine so that the radioactivity present in the initial HCl–NaCl extract (i.e., [14C]leucine) increased as that in the final NaOH extract (i.e., 14C-labeled protein) decreased. Using this new assay for cell protein, free and immobilized cell cultures were found to exhibit virtually identical kinetics of glucose utilization, growth, and patulin production. In addition to providing a means of comparing the specific productivity of free versus immobilized cell preparations, this assay accurately measures the incorporation of [14C]leucine into cellular protein and could be used as a measure of cell viability.

The Effect of Dilution Rate upon Protein Content and Cellular Amino Acid Profiles in Chemostat Cultures of Saccharomyces Cerevisiae CABI 039916

2010 ◽  
Vol 6 (4) ◽  
Author(s):  
Beverley Finn ◽  
Linda M Harvey ◽  
Brian McNeil
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Protein Content ◽  
Dilution Rate ◽  
Specific Growth ◽  
Cellular Protein ◽  
Cell Protein ◽  
Protein Pool ◽  
Cellular Protein Content ◽  
Amino Acid Profiles

In the present study we examined the use of a chemostat system to investigate the impact of changes in the specific growth rate of Saccharomyces cerevisiae CABI 039916 on cellular amino acid profiles and total protein content. This experimental system allowed the unambiguous examination of the link between changes in dilution rate and the culture response, which would have been difficult in batch or fed-batch cultures. Alteration of the specific growth rate (via manipulation of the dilution rate) within a carbon and energy-limited chemostat has a significant impact on the physiology of Saccharomyces cerevisiae. Low dilution rates (<0.1h-1) led to predominantly respiratory metabolism and the maximisation of cellular protein content within the cell (58%), by contrast high dilution rates (>0.2h-1) led to respirofermentative metabolism, where the cellular protein content was minimal (~40%). The content of nearly all amino acids in the yeast protein pool fell significantly as dilution rate increased in parallel with the decline in cell protein content. By contrast, the concentration of two related key food/feed amino acids in the cell protein pool—glutamic acid and arginine could be increased within the cellular protein by 5% (increasing the dilution rate from 0.05h-1 to 0.25h-1) and 1.5% respectively (decreasing the dilution rate from 0.05h-1 to 0.2h-1). Despite previous studies showing that metabolic change was associated with major changes in free amino acid levels, the present study indicates that the total cellular yeast protein amino acid composition is largely invariant despite profound metabolic changes, with one or two key exceptions.

ATP and Protein Synthesis Assays to Evaluate the Toxicities of Preservatives in Vitro

1991 ◽  
Vol 19 (1) ◽  
pp. 60-67
Author(s):  
Corrado L. Galli ◽  
Barbara Viviani ◽  
Giampiero Casartelli ◽  
Marina Marinovich
Keyword(s):  
Protein Synthesis ◽  
Protein Content ◽  
Cellular Protein ◽  
Cell Protein ◽  
Leucine Incorporation ◽  
Maximal Effect ◽  
Atp Level ◽  
Cellular Protein Content

Cellular protein content, protein synthesis, ATP level and lactate dehydrogenase (LDH) activity, measured in a murine epidermal cell line (HEL/30), were used as the endpoints for determining the cytotoxicities of 17 antimicrobial chemicals. The relative toxicities of the test compounds were quantified by the determination of the concentrations inducing a 50% inhibition of [3H]-leucine incorporation into proteins (IC50), causing a 50% reduction of ATP level or final cell protein content or producing the maximal effect on LDH leakage (EC50) after 2 hours of treatment. The results indicate a good correlation between both the reduction of ATP level and inhibition of protein synthesis and the minimal inhibitory concentration (MIC) on different microorganisms, suggesting that ATP and protein synthesis assays could be useful as prescreening methods for testing the cytotoxicities of preservatives.

scholarly journals Tubular cell protein degradation in early diabetic renal hypertrophy.

1994 ◽  
Vol 4 (8) ◽  
pp. 1582-1587
Author(s):  
P Shechter ◽  
G Boner ◽  
R Rabkin
Keyword(s):  
Protein Degradation ◽  
Protein Content ◽  
Cellular Protein ◽  
Diabetic Rats ◽  
Tubular Cell ◽  
Cell Protein ◽  
Renal Hypertrophy ◽  
Cathepsin Activity ◽  
Wet Weight

Renal hypertrophy in diabetes is accompanied by an increase in kidney protein content, which reflects an imbalance between protein synthesis and degradation. This study determines whether altered cellular protein degradation contributes to the imbalance. Diabetes was induced in rats with streptozotocin (55 mg/kg/ip). After 2 or 4 days of diabetes, kidney weight and protein content were measured. Over the 4 days, despite a loss in body weight, kidney wet weight increased by 35% and protein content by 37% in the diabetic rats. Treatment with insulin prevented this increase. Long-lived protein degradation was measured in isolated proximal tubules prelabeled with (14C)valine in vivo. Two days after streptozotocin, protein degradation was depressed by 19% (P < 0.05) and by the fourth day by 27% compared with that in nondiabetic controls (2.6% +/- 0.2 versus 1.9 +/- 0.1% degraded/h; P < 0.01). This was accompanied by a similar diabetes-induced decrease in proximal tubule cathepsin B and L activity. Accordingly, this study provides direct evidence that, in diabetes, tubular cell protein breakdown is depressed and suggests that altered lysosomal cathepsin activity may contribute to this effect. Depressed proteolysis likely contributes to the increase in kidney protein content and hence to diabetic renal hypertrophy.

Design of a continuous flow immobilized-cell reactor for biosynthetical production of L-aspartic acid

1985 ◽  
Vol 50 (10) ◽  
pp. 2122-2133 ◽  
Author(s):  
Jindřich Zahradník ◽  
Marie Fialová ◽  
Jan Škoda ◽  
Helena Škodová
Keyword(s):  
Aspartic Acid ◽  
Continuous Flow ◽  
Immobilized Cells ◽  
Scale Up ◽  
Fixed Bed ◽  
Packed Bed ◽  
Fixed Bed Reactor ◽  
Experimental Program ◽  
Design Parameters ◽  
Immobilized Cell

An experimental study was carried out aimed at establishing a data base for an optimum design of a continuous flow fixed-bed reactor for biotransformation of ammonium fumarate to L-aspartic acid catalyzed by immobilized cells of the strain Escherichia alcalescens dispar group. The experimental program included studies of the effect of reactor geometry, catalytic particle size, and packed bed arrangement on reactor hydrodynamics and on the rate of substrate conversion. An expression for the effective reaction rate was derived including the effect of mass transfer and conditions of the safe conversion-data scale-up were defined. Suggestions for the design of a pilot plant reactor (100 t/year) were formulated and decisive design parameters of such reactor were estimated for several variants of problem formulation.

Prediction of Human Acute Toxicity by the Hep G2/24-hour/Total Protein Assay, with Protein Measurement by the CBQCA Method

2005 ◽  
Vol 33 (3) ◽  
pp. 207-213 ◽  
Author(s):  
Paul J. Dierickx
Keyword(s):  
Protein Content ◽  
Total Protein ◽  
Total Protein Content ◽  
Human Toxicity ◽  
Hep G2 ◽  
Vitro Assay ◽  
Protein Assay ◽  
Lowry Method ◽  
Total Protein Assay

In our previously described Hep G2/24-hour/total protein assay, protein levels were measured by using the Lowry method. This assay was the best acute in vitro assay for the prediction of human toxicity within the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) study. In order to increase the MEIC data-base with a wider range of chemicals, we were interested in introducing the more practical 3-(4-carboxybenzoyl)-quinoline-2-carboxaldehyde (CBQCA) method for the quantification of the total protein content. Therefore, we investigated whether the same good results for the prediction of acute human toxicity would be obtained with the CBQCA method. The cells were treated for 24 hours, then cytotoxicity was determined by measuring the total protein content with CBQCA. The results were quantified by using the PI50c: the concentration (in mM) of test compound required to reduce the total protein content measured with the CBQCA-method by 50% as compared to the control cells. The results were compared with the PI50, the corresponding value when the Lowry method was used. A relatively low correlation was observed between PI50 and PI50c, reflecting the large and unexpected, differences when using the two protein assays. However, when comparing the log PI50c with the human toxicity, a correlation coefficient of r2 = 0.761 ( n = 44) was obtained for exactly the same series of MEIC chemicals. This value is clearly higher than that for the Lowry method ( r2 = 0.695). Compared to the Lowry method originally used, the Hep G2/24-hour/CBQCA total protein assay has the additional important advantage that it can be very easily adapted for large-scale analyses with robotic systems, including the on-line calculation of the results.

Ekstraksi Protein dari Rhodophyta dan Chlorophyta Dari Perairan Pulau Pari Sebagai Alternatif Antioksidant

2020 ◽  
Vol 1 (1) ◽  
pp. 24-31
Author(s):  
Elvi Kustiyah ◽  
Bungaran Saingin ◽  
Hernowo Widodo ◽  
Viriya Piti
Keyword(s):  
Antioxidant Activity ◽  
Low Temperature ◽  
Protein Content ◽  
Green Algae ◽  
Red Algae ◽  
Marine Resources ◽  
Activity Test ◽  
Ic50 Value ◽  
Dpph Method ◽  
Lowry Method

 Indonesia has millions island and big part of Indonesia is sea that is rich in marine biological resources and has the potential to be developed and optimized. One of the abundant marine resources in Indonesia is algae. Algae are plant-like protists. Algae have several important compounds, including protein, carbohydrates, fats, minerals and other useful elements. Proteins contained in algae have the potential to be used as antioxidants. In this study, the levels of protein in red and green algae were tested by using the lowry method and testing the antioxidant activity of red and green algae extracts using the Diphenylpicrylhydrazyl (DPPH) method. Algae extraction was done by maceration, which is soaking the sample in low temperature with phosphate buffer saline (PBS) pH 7. From the extraction results it can be concluded that the red algae (Rhodophyta) has the highest protein content of 5.115 ± 0.126% and the lowest protein content in green algae (Chloropytha) as big as 1.686 ± 0.430%. And from the results of the antioxidant activity test showed that all positive algae showed antioxidant activity but the green algae (Chlorophyta) had the highest antioxidant activity of 71.5946 ± 0.01612% with IC50 value 1.6114.

scholarly journals The same normal cell protein is phosphorylated after transformation by avian sarcoma viruses with unrelated transforming genes.

1981 ◽  
Vol 1 (1) ◽  
pp. 43-50 ◽  
Author(s):  
E Erikson ◽  
R Cook ◽  
G J Miller ◽  
R L Erikson
Keyword(s):  
Rous Sarcoma Virus ◽  
Direct Consequence ◽  
Cellular Protein ◽  
Cell Protein ◽  
Rous Sarcoma ◽  
Sarcoma Virus ◽  
Transforming Proteins ◽  
Avian Sarcoma ◽  
Transforming Genes ◽  
Chicken Embryo Fibroblasts

The phosphorylation of a normal cellular protein of molecular weight 34,000 (34K) is enhanced in Rous sarcoma virus-transformed chicken embryo fibroblasts apparently as a direct consequence of the phosphotransferase activity of the Rous sarcoma virus-transforming protein pp60src. We have prepared anti-34K serum by using 34K purified from normal fibroblasts to confirm that the transformation-specific phosphorylation described previously occurs on a normal cellular protein and to further characterize the nature of the protein. In this communication, we also show that the phosphorylation of 34K is also increased in cells transformed by either Fujinami or PRCII sarcoma virus, two recently characterized avian sarcoma viruses whose transforming proteins, although distinct from pp60src, are also associated with phosphotransferase activity. Moreover, comparative fingerprinting of tryptic phosphopeptides shows that the major site of phosphorylation of 34K is the same in all three cases.

Bioethanol Production by Pichia stipitis Immobilized on Water Hyacinth and Thin-Shell Silk Cocoon

2021 ◽  
Author(s):  
Suchata Kirdponpattara ◽  
Santi Chuetor ◽  
Malinee Sriariyanun ◽  
Muenduen Phisalaphong
Keyword(s):  
Water Hyacinth ◽  
Thin Shell ◽  
Immobilized Cells ◽  
High Surface ◽  
Pichia Stipitis ◽  
High Porosity ◽  
Bioethanol Production ◽  
Immobilized Cell ◽  
Silk Cocoon ◽  
Cell Carriers

Cell immobilization technique was applied in this study in order to examine effect of immobilized Pichia stipitis TISTR5806 on bioethanol production. Water hyacinth (WH) and thin-shell silk cocoon (CC) were used as cell carriers. Characteristics of the cell carriers were examined to explain the mechanism of bioethanol production. Carrier sizes and weights were optimized to improve bioethanol production. Moreover, stabilities of immobilized cells and carriers were evaluated. Because of high porosity, high surface area and good swelling ability of WH, cell immobilized on 1 g WH with 1 cm length produced the highest ethanol concentration at 13.3 g/L. Five cycles of a repeated batch of immobilized cell (IC) system on WH showed stable performance in ethanol production (8.2–10.4 g/L) with large numbers of the immobilized cells. The interaction between the immobilized cells and the WH surface were discovered.

Effect of adenovirus on metabolism of specific host mRNAs: transport control and specific translational discrimination

1983 ◽  
Vol 3 (7) ◽  
pp. 1212-1221 ◽  
Author(s):  
A Babich ◽  
L T Feldman ◽  
J R Nevins ◽  
J E Darnell ◽  
C Weinberger
Keyword(s):  
Protein Synthesis ◽  
Host Cell ◽  
Cellular Protein ◽  
Host Protein ◽  
Cell Protein ◽  
Viral Mrna ◽  
Initiation Of Translation ◽  
Viral Mutant ◽  
Cellular Mrnas ◽  
Cellular Protein Synthesis

We have studied the adenovirus-induced inhibition of host cell protein synthesis and the effect of infection on the overall metabolism of host cell mRNA during the late phase of adenovirus infection by following the fate of a number of cellular mRNAs complementary to specific cloned DNA segments. At a time in infection when the rate of total cellular protein synthesis is drastically (greater than 90%) reduced, transcription of specific cellular genes is undiminished. However, the transport of newly synthesized cellular mRNA to the cytoplasm is greatly decreased. This decreased appearance of new mRNA in the cytoplasm cannot account for the observed cessation of cell specific protein synthesis, however, since the concentration of several preexisting cellular mRNAs, including the mRNA for actin, remains unchanged throughout the course of infection. The preexisting mRNA is intact, capped, and functional as judged by its ability to direct protein synthesis in vitro in a cap-dependent fashion. The interruption in host translation appears to operate at the level of initiation directly, since we find that fewer ribosomes are associated with a given cellular mRNA after infection than before infection. Furthermore, the in vivo inhibition of cellular protein synthesis does not appear to be the result of competition with viral mRNA, since conditions which prevent the efficient initiation of translation of viral mRNA (infection with a viral mutant) do not result in the recovery of cell translation. Thus, it appears that a late adenovirus gene product directly mediates a shutoff of host protein synthesis.

Effect of Tacca Starch Loading on Production of Amylolytic Enzymes from Ragi Tapai

2020 ◽  
Vol 987 ◽  
pp. 118-123
Author(s):  
Nurul Syafiqa Izyan ◽  
Dg Nurdayana Azman ◽  
Nur Amalina Mohd Saad ◽  
Suhaila Mohd Sauid ◽  
Fazlena Hamzah
Keyword(s):  
Protein Content ◽  
Total Protein ◽  
Amylase Activity ◽  
Total Protein Content ◽  
Growth Promoters ◽  
Amylolytic Enzyme ◽  
Amylolytic Enzymes ◽  
Lowry Method ◽  
Medium Increase ◽  
Optimum Production

The study was done to determine the effect of Tacca starch loading on production of amylolytic enzyme from Ragi Tapai. In this study, Ragi Tapai was used as a starter to produce amylolytic enzyme. The fermentation was done in a solid state fermentation with the presence of Tacca leontopetaloides starch as the carbon source. The analysis of total sugar was conducted using DNS method and amylolytic enzyme was determined using Lowry method. The mixture was fermented and incubated for 24, 48, 72 and 96h. The result revealed that the optimum production of amylase was found at 48 h of incubation with amylase activity of 1.91 U/ml/min and 1.42 mg/ml for total protein. The study shows that increment amount of the Tacca starch in cultivation medium, increase the production of the amylase and total protein content. The highest enzyme activity was obtained at 4% of Tacca starch loading with amylase activity and total protein content of 2.14 U/ml/min and 1.42 mg/ml respectively. The study indicated that growth promoters in Tacca starch capable to enhance the activity of microbial consortium in Ragi Tapai for production of the amylolytic enzyme.

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