Untersuchungen über den Kaliumtransport in E. coli B

1971 ◽  
Vol 26 (9) ◽  
pp. 887-893 ◽  
Author(s):  
U. Zimmermann ◽  
G. Pilwat
Keyword(s):  
Amino Acids ◽  
Incubation Medium ◽  
Potassium Level ◽  
Potassium Concentration ◽  
Essential Amino Acids ◽  
Type B ◽  
Wild Type ◽  
E Coli ◽  
Stabilization Effect ◽  
Intracellular Potassium Concentration

An intracellular concentration of potassium in E. coli B 525 stationary for several hours is only attained by addition of the essential amino acids histidine and leucine, or ammonium chloride, to the incubation medium. The level of the stationary intracellular potassium concentration is dependent upon the concentration of these substances.The two other combinations of the essential amino acids (leucine/methionine and histidine/methionine) and the single amino acids show no stabilization effect upon the internal stationary potassium level.In the case of the wild type B 163, the instability of the stationary internal potassium concentration, and the corresponding effect of the above mentioned substances, is not so significant. Therefore the instability which is observed in the case of B 525 can be deduced from the mutation.

scholarly journals Karakteristik Fisiko-Kimia Konsentrat Protein Ikan Sunglir (Elagatis bipinnulatus)

Jurnal MIPA ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 164
Author(s):  
Frets Jonas Rieuwpassa ◽  
Eko Cahyono
Keyword(s):  
Amino Acids ◽  
Physicochemical Properties ◽  
Essential Amino Acids ◽  
Fat Content ◽  
Pelagic Fish ◽  
Animal Protein ◽  
Physical Analysis ◽  
Fat Absorption ◽  
Type B ◽  
High Level

Ikan merupakan sumber protein hewani yang memiliki daya cerna yang lebih baik dan jumlah kandungan asam amino essensial yang lebih banyak dibandingkan dengan sumber protein hewani lainnya. Ikan sunglir adalah jenis ikan pelagis yang banyak hidup diperairan Nusa Utara. Ekstraksi KPI umumnya dilakukan dengan menggunakan pelarut etanol. Penelitian ini bertujuan mengkarakterisasi sifat fisiko-kimia konsentrat protein ikan yang diekstrak dari ikan sunglir. Penggunaan etanol 90% dalam mengekstraksi konsentrat protein dari ikan sunglir menghasilkan rendemen berkisar 18-20%. Hasil Penelitian menunjukkan bahwa konsentrat protein ikan sunglir memiliki kandungan protein yang tinggi dan memiliki kadar lemak yang rendah. Konsentrat protein dengan kadar protein >65% dan kadar lemak <3% tergolong sebagai konsentrat protein Tipe B sesuai dengan standar Mutu FAO 1976 tentang KPI. Hasil pengujian fisik menunjukkan bahwa KPI memiliki kemampuan penyerapan air, lemak dan densitas kamba yang cukup baik untuk diaplikasikan ke dalam bahan panganFish serve as an important source of animal protein with better digestibility and higher content of essential amino acids than other sources of animal protein. Elagatis bipinnulatus or sunglir in Indonesian is a common pelagic fish caught in Sangihe Islands. FPC is commonly extracted with etanol. Therefore, this research aims to characterise the physicochemical properties of the FPC extracted from rainbow runner. The use of 90% ethanol for exraction of FPC from rainbow runner resulted in 18-20% yield. The result shows that the local rainbow runner contained FPC with high level of protein (77.34%) but low level of fat (1.22%), classified as type B on the basis of FAO’s standard on FPC Protein, which stipulated FPC with >65% protein and <3% fat content as Type B. In addition, physical analysis proved that FPC has appropriate water and fat absorption abilities as well as kamba density, suitble for food substitute or fortification

The Bβ-sheet in the PAI-1 Molecule Plays an Important Role for Its Stability

2000 ◽  
Vol 83 (06) ◽  
pp. 896-901 ◽  
Author(s):  
Guang-Chao Sui ◽  
Björn Wiman
Keyword(s):  
Amino Acids ◽  
Half Life ◽  
Ph Dependence ◽  
The Other ◽  
Wild Type ◽  
E Coli ◽  
Reactive Center ◽  
The Stability ◽  
Β Sheet ◽  
Pai 1

SummaryWe have investigated the B β-sheet in PAI-1 regarding its role for the stability of the molecule. The residues from His219 to Tyr241 (except for Gly230 and Pro240), covering the s2B and s3B strands, and in addition His185 and His190 were substituted by amino acids with opposite properties. The 23 generated single-site changed mutants and also wild type PAI-1 (wtPAI-1) were expressed in E. coli. Subsequently they were purified by heparin-Sepharose and anhydrotrypsin agarose affinity chromatographies. The stability of the purified PAI-1 variants was analyzed at 37° C and at different pHs (5.5, 6.5 or 7.5). At pH 7.5 and 37° C, single substitutions of the residues in the central portions of both strands 2 and 3 in the B β-sheet (Ile223 to Leu226 on s2B and Met235 to Ile237 on s3B), caused a significant decrease in stability, yielding half-lives of about 10–25% as compared to wtPAI-1. On the other hand, mutations at both sides of the central portion of the B β-sheet (Tyr221, Asp222, Tyr228 and Thr232) frequently resulted in an increased PAI-1 stability (up to 7-fold). While wtPAI-1 exhibited prolonged half-lives at pH 6.5 and 5.5, the PAI-1 variant Y228S was more stable at neutral pH (half-life of 9.6 h at pH 7.5) as compared to its half-life at pH 5.5 (1.1 h). One of the 4 modified histidine residues (His229) resulted in a variant with a clearly affected stability as a function of pH, suggesting that it may, at least in part, be of importance for the pH dependence of the PAI-1 stability. Thus, our data demonstrate that the B β-sheet is of great importance for the stability of the molecule. Modifications in this part causes decreased or increased stability in a certain pattern, suggesting effects on the insertion rate of the reactive center loop into the A β-sheet of the molecule.

Molecular Characterization of Genes Coding for Wild-Type and Sulfonylurea-Resistant Acetolactate Synthase in the Cyanobacterium Synechococcus PC C7942

1990 ◽  
Vol 45 (5) ◽  
pp. 538-543 ◽  
Author(s):  
D. Friedberg ◽  
J. Seijffers
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Characterization ◽  
Amino Acid Level ◽  
Acetolactate Synthase ◽  
Mutant Gene ◽  
Wild Type ◽  
E Coli

We present here the isolation and molecular characterization of acetolactate synthase (ALS) genes from the cyanobacterium Synechococcus PCC7942 which specify a sulfonylurea-sensitive enzyme and from the sulfonylurea-resistant mutant SM3/20, which specify resistance to sulfonylurea herbicides. The ALS gene was cloned and mapped by complementation of an Escherichia coli ilv auxotroph that requires branched-chain amino acids for growth and lacks ALS activity. The cyanobacterial gene is efficiently expressed in this heterologous host. The ALS gene codes for 612 amino acids and shows high sequence homology (46%) at the amino acid level with ALS III of E. coli and with the tobacco ALS. The resistant phenotype is a consequence of proline to serine substitution in residue 115 of the deduced amino acid sequence. Functional expression of the mutant gene in wild-type Synechococcus and in E. coli confirmed that this amino-acid substitution is responsible for the resistance. Yet the deduced amino-acid sequence as compared with othjer ALS proteins supports the notion that the amino-acid context of the substitution is important for the resistance.

THE STIMULATION OF INSULIN RELEASE BY ESSENTIAL AMINO ACIDS FROM RABBIT PANCREAS IN VITRO

1970 ◽  
Vol 47 (3) ◽  
pp. 347-356 ◽  
Author(s):  
R. D. G. MILNER
Keyword(s):  
Amino Acids ◽  
Insulin Release ◽  
Incubation Medium ◽  
Statistical Significance ◽  
Adenosine Monophosphate ◽  
Essential Amino Acids ◽  
Isoleucine Leucine ◽  
Rabbit Pancreas

SUMMARY Pieces of rabbit pancreas were incubated in vitro in an incubation medium containing no glucose or 1·5 mg. glucose/ml. In each of these conditions the effect on insulin release of each of the essential amino acids at 5 mm concentration was studied. Leucine was the only essential amino acid that stimulated insulin release to a level which reached statistical significance in an incubation medium containing no glucose. In medium containing 1·5 mg. glucose/ml., arginine, isoleucine, leucine and lysine stimulated insulin release and phenylalanine inhibited insulin release. Glucagon, theophylline or dibutyryl cyclic adenosine monophosphate stimulated insulin release significantly in the presence of leucine but not in the presence of arginine. Arginine stimulated insulin release in the presence of leucine. The results of these experiments characterize further the difference in the mechanism of action of leucine and arginine on the pancreatic β-cell and indicate possible explanations for results obtained in other species in vivo.

scholarly journals Chemotaxis of Escherichia coli to Pyrimidines: a New Role for the Signal Transducer Tap

2007 ◽  
Vol 190 (3) ◽  
pp. 972-979 ◽  
Author(s):  
Xianxian Liu ◽  
Rebecca E. Parales
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Growth Conditions ◽  
Wild Type ◽  
E Coli ◽  
C Terminus ◽  
Signaling Domain ◽  
Chemotaxis Proteins ◽  
Periplasmic Domain

ABSTRACT Escherichia coli exhibits chemotactic responses to sugars, amino acids, and dipeptides, and the responses are mediated by methyl-accepting chemotaxis proteins (MCPs). Using capillary assays, we demonstrated that Escherichia coli RP437 is attracted to the pyrimidines thymine and uracil and the response was constitutively expressed under all tested growth conditions. All MCP mutants lacking the MCP Tap protein showed no response to pyrimidines, suggesting that Tap, which is known to mediate dipeptide chemotaxis, is required for pyrimidine chemotaxis. In order to confirm the role of Tap in pyrimidine chemotaxis, we constructed chimeric chemoreceptors (Tapsr and Tsrap), in which the periplasmic and cytoplasmic domains of Tap and Tsr were switched. When Tapsr and Tsrap were individually expressed in an E. coli strain lacking all four native MCPs, Tapsr mediated chemotaxis toward pyrimidines and dipeptides, but Tsrap did not complement the chemotaxis defect. The addition of the C-terminal 19 amino acids from Tsr to the C terminus of Tsrap resulted in a functional chemoreceptor that mediated chemotaxis to serine but not pyrimidines or dipeptides. These results indicate that the periplasmic domain of Tap is responsible for detecting pyrimidines and the Tsr signaling domain confers on Tapsr the ability to mediate efficient chemotaxis. A mutant lacking dipeptide binding protein (DBP) was wild type for pyrimidine taxis, indicating that DBP, which is the primary chemoreceptor for dipeptides, is not responsible for detecting pyrimidines. It is not yet known whether Tap detects pyrimidines directly or via an additional chemoreceptor protein.

scholarly journals Engineered Cyanophycin Synthetase (CphA) from Nostoc ellipsosporum Confers Enhanced CphA Activity and Cyanophycin Accumulation to Escherichia coli

2006 ◽  
Vol 72 (12) ◽  
pp. 7652-7660 ◽  
Author(s):  
Tran Hai ◽  
Kay M. Frey ◽  
Alexander Steinbüchel
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Dry Matter ◽  
Wild Type ◽  
Acinetobacter Baylyi ◽  
E Coli ◽  
Recombinant Cells ◽  
Hydrophobic Amino Acids ◽  
Type Gene ◽  
Nostoc Ellipsosporum

ABSTRACT The cyanophycin (CGP) synthetase gene (cphA NE1) of the transposon-induced argL mutant NE1 of the cyanobacterium Nostoc ellipsosporum, which exhibits a CGP-leaky phenotype during diazotrophical growth, was cloned and expressed in Escherichia coli strain TOP10. Its amino acid sequence exhibited high similarities to CphAs of other cyanobacteria. Recombinant cells of E. coli, which harbored a fragment comprising the complete cphA NE1 gene plus 400 bp of its downstream region in colinear orientation to the lacZ promoter, accumulated CGP up to 17 and 8.5% (wt/wt) of cellular dry matter (CDM) if cultivated in complex medium in the presence or absence of isopropyl-β-d-thiogalactopyranoside, respectively. Two truncated CphAs, lacking 31 (CphANE1del96) or 59 (CphANE1del180) amino acids of the C-terminal region, were derived from cphA NE1 by deleting 96 or 180 bp from its 3′ region through the introduction of stop codons. In comparison to the wild-type gene, cphA NE1del96 conferred about 2.1- to 2.2-fold-higher enzyme activity (up to 5.75 U/mg protein) on E. coli. Furthermore, these cells accumulated about twofold more CGP (up to 34.5% [wt/wt] of CDM) than cells expressing the wild-type gene. An engineered CphA possessing significantly enhanced activity and conferring the highest CGP content on E. coli is demonstrated. In contrast, CphANE1del180 was inactive and did not confer CGP accumulation on E. coli. Interestingly, a short conserved stretch of 4 to 5 hydrophobic amino acids is located in the protein region present in CphANE1del96 but absent in CphANE1del180. In addition, CphANE1 and CphANE1del96 are, besides CphA from Acinetobacter baylyi, the only CphAs exhibiting rigid substrate specificities that do not enable the incorporation of lysine instead of arginine into CGP.

Accumulation of casein-derived peptides during growth of proteinase-positive strains of Lactococcus lactis in milk: their contribution to subsequent bacterial growth is impaired by their internal transport

2000 ◽  
Vol 67 (2) ◽  
pp. 233-240 ◽  
Author(s):  
CATHERINE FOUCAUD ◽  
VINCENT JUILLARD
Keyword(s):  
Amino Acids ◽  
Lactococcus Lactis ◽  
Nutritional Value ◽  
Essential Amino Acids ◽  
Peptide Transport ◽  
Wild Type ◽  
Transport Mutants ◽  
Time Courses ◽  
Internal Transport ◽  
Type Strains

To explain the limited nutritional value of milk cultured with proteinase-positive (Prt+) strains of Lactococcus lactis for the subsequent growth of dairy lactococci, we investigated further the time courses of modifications in the free amino acid and peptide contents of cultured milk. When growing in milk for up to 24 h, Prt+ strains of Lc. lactis progressively accumulated amino acids and casein-derived peptides. The growth of proteinase-negative (Prt−) wild-type strains and peptide transport mutants of Lc. lactis in cultured milk showed that casein-derived peptides could sustain growth up to 5×108 cfu/ml, depending on the extent of casein degradation during the preliminary growth of Prt+ strains and the Prt− strains. Of the casein-derived oligopeptides, <25% were transported into the cell and used for Lc. lactis growth. However, they played a prominent role, contributing 90% to growth. In contrast, di- and tripeptides did not contribute to growth, suggesting that either few were released from caseins or they did not supply essential amino acids.

scholarly journals Members of the Conserved DedA Family Are Likely Membrane Transporters and Are Required for Drug Resistance in Escherichia coli

2013 ◽  
Vol 58 (2) ◽  
pp. 923-930 ◽  
Author(s):  
Sujeet Kumar ◽  
William T. Doerrler
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Drug Resistance ◽  
Public Health Problem ◽  
Membrane Transporters ◽  
Wild Type ◽  
Content Type ◽  
E Coli ◽  
Acidic Amino Acids ◽  
Genes Encoding

ABSTRACTBacterial resistance to antibiotics and biocides is an increasing public health problem. Genes encoding integral membrane proteins belonging to the DedA family are present in most bacterial genomes, includingEscherichia coli. AnE. colistrain lacking partially redundant DedA family genesyqjAandyghB(strain BC202) displays temperature sensitivity and cell division defects. These phenotypes can be corrected by overexpression ofmdfA, an Na+-K+/H+antiporter of the major facilitator superfamily. We show that BC202 is hypersensitive to several biocides and cationic compounds that are known substrates of several multidrug resistance transporters, including MdfA, EmrE, and AcrB. The introduction of deletions of genes encoding these drug transporters into BC202 results in additional sensitivity. Expression of wild-typeyghBoryqjAcan restore drug resistance, but this is eliminated upon mutation of two membrane-embedded acidic amino acids (E39 or D51 in either protein). This dependence upon membrane-embedded acidic amino acids is a hallmark of proton-dependent antiporters. Overexpression ofmdfAin BC202 or artificially restoring proton motive force (PMF) restores wild-type resistance to substrates of MdfA as well as other drug resistance transporters such as EmrE and AcrAB. These results suggest that YqjA and YghB may be membrane transporters required for PMF-dependent drug efflux inE. coli.

scholarly journals Functional Analysis of an Essential GSP1/Ran Ortholog Gene, CpRan1, from the Chestnut Blight Fungus Cryphonectria parasitica Using a Heterokaryon

2021 ◽  
Vol 7 (5) ◽  
pp. 332
Author(s):  
Dae-Hyuk Kim ◽  
Yo-Han Ko ◽  
Jeesun Chun
Keyword(s):  
Amino Acids ◽  
Functional Analysis ◽  
Mutant Allele ◽  
Microscopic Analysis ◽  
Essential Amino Acids ◽  
Cryphonectria Parasitica ◽  
Complementation Analysis ◽  
Null Mutant ◽  
Wild Type ◽  
Chestnut Blight Fungus

Functional analysis of a GSP1/Ran ortholog, CpRan1, from Cryphonectria parasitica was conducted. Genotype analysis revealed that the putative CpRan1-null mutant was a heterokaryotic transformant harboring two different types of nuclei, one with the wild-type CpRan1 allele and the other with the CpRan1-null mutant allele. The mycelial growth and colony morphology of the heterokaryotic transformant was normal. Microscopic analysis of the resulting conidia (aseptate and monokaryotic asexual spores) demonstrated that although normal germinating spores were observed from conidia harboring a nucleus with the wild-type CpRan1 allele, a number of residual conidia that did not germinate existed. Complementation analysis using protoplasts from the heterokaryon with the wild-type CpRan1 allele confirmed that the CpRan1 gene is essential to C. parasitica. Complementation analysis using the various CpRan1 chimera constructs allowed us to perform a functional analysis of essential amino acids of the CpRan1. Among the four suggested essential amino acids, Lys-97 for ubiquitination was determined to not be an essential residue. Moreover, the CpRan1-null mutant allele was successfully complemented with mouse Ran gene, which suggested that the biological function of Ran gene is evolutionary conserved and that our heterokaryon rescue can be applied for the functional analysis of heterologous genes.

scholarly journals Communicating the nutritional value of sugar in Drosophila

2018 ◽  
Vol 115 (12) ◽  
pp. E2829-E2838 ◽  
Author(s):  
Farhan Abu ◽  
Justin G. Wang ◽  
Yangkyun Oh ◽  
Jingjing Deng ◽  
Thomas A. Neubert ◽  
...  
Keyword(s):  
Amino Acids ◽  
Fatty Acids ◽  
Real Time ◽  
Olfactory System ◽  
Nutritional Value ◽  
Essential Amino Acids ◽  
Wild Type ◽  
Correlation Analyses

Sweet-insensitive Drosophila mutants are unable to readily identify sugar. In presence of wild-type (WT) flies, however, these mutant flies demonstrated a marked increase in their preference for nutritive sugar. Real-time recordings of starved WT flies revealed that these flies discharge a drop from their gut end after consuming nutritive sugars, but not nonnutritive sugars. We proposed that the drop may contain a molecule(s) named calorie-induced secreted factor (CIF), which serves as a signal to inform other flies about its nutritional value. Consistent with this, we observed a robust preference of flies for nutritive sugar containing CIF over nutritive sugar without CIF. Feeding appears to be a prerequisite for the release of CIF, given that fed flies did not produce it. Additionally, correlation analyses and pharmacological approaches suggest that the nutritional value, rather than the taste, of the consumed sugar correlates strongly with the amount (or intensity) of the released CIF. We observed that the release of this attractant signal requires the consumption of macronutrients, specifically nutritive sugars and l-enantiomer essential amino acids (l-eAAs), but it is negligibly released when flies are fed nonnutritive sugars, unnatural d-enantiomer essential amino acids (d-eAAs), fatty acids, alcohol, or salts. Finally, CIF (i) is not detected by the olfactory system, (ii) is not influenced by the sex of the fly, and (iii) is not limited to one species of Drosophila.

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