scholarly journals Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. The involvement of sulphur metabolism

1973 ◽  
Vol 136 (3) ◽  
pp. 477-490 ◽  
Author(s):  
Albert Neuberger ◽  
John D. Sandy ◽  
George H. Tait
Keyword(s):  
De Novo ◽  
Maximum Activity ◽  
Fresh Medium ◽  
Synthetase Activity ◽  
Sulphur Metabolism ◽  
Sulphur Compounds ◽  
Similar Treatment ◽  
Rhodopseudomonas Spheroides ◽  
Low Activity

1. The ‘initial’ 5-aminolaevulinate synthetase activity, that is the activity observed immediately after cell disruption, in extracts prepared from unharvested semianaerobically grown Rhodopseudomonas spheroides, was twice that observed under the same assay conditions in extracts prepared from harvested cells. 2. The effect of oxygenation of a culture on the ‘maximum’ aminolaevulinate synthetase activity, that is the activity observed 1h after disruption of harvested cells, is markedly influenced by the contents of the growth medium. Oxygenation of organisms for 1h in the medium in which they have grown produces an 80–90% decrease in maximum activity, whereas similar treatment of organisms resuspended in fresh medium produces less than a 40% decrease. 3. This protective effect of fresh medium is absolutely dependent on the presence of sulphate. When cells are suspended in sulphate-deficient fresh medium, the maximum activity falls by 65–75% even without oxygenation. A high maximum activity is regenerated when sulphate is resupplied. 4. When organisms are oxygenated in the medium in which they have grown, the cellular contents of GSH+GSSG and cysteine+cystine fall very markedly and homolanthionine is formed. Both the fall in aminolaevulinate synthetase activity and the changes in sulphur metabolism are largely prevented by the addition of compounds which stimulate synthesis of cysteine de novo or inhibit the conversion of cysteine S into homocysteine S. 5. The maximum aminolaevulinate synthetase activity was directly proportional to the GSH+GSSG content of all cell preparations. In glutathione-depleted extracts the ‘low’-activity enzyme could be re-activated in vitro by the addition of GSH, GSSG, cysteine or cystine, whereas in extracts with a high glutathione content the ‘high’-activity enzyme was unaffected by these sulphur compounds. 6. The activation of low-activity enzyme with exogenous sulphur compounds was prevented by excluding air or by adding NADH. Studies with purified enzyme indicate that sulphur compounds do not interact directly with the enzyme, but that their effect is mediated by a number of other endogenous factors.

scholarly journals Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides a role for trisulphides

1975 ◽  
Vol 150 (2) ◽  
pp. 245-257 ◽  
Author(s):  
J D Sandy ◽  
R C Davies ◽  
A Neuberger
Keyword(s):  
Ion Exchange ◽  
Extraction Procedure ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Synthetase Activity ◽  
Multiple Forms ◽  
Anaerobic Culture ◽  
Structural Requirement ◽  
Rhodopseudomonas Spheroides ◽  
Low Activity

1. The aminolaevulinate synthetase activator of fresh extracts of semi-anaerobically grown Rhodopseudomonas spheroids was resolved into two fractions by ion-exchange chromatography. One fraction was identified as cystine trisulphide (CySSSCy). Analysis of the other fraction indicated the presence of about equal amounts of glutathione trisulphide (GSSSG) and the mixed trisulphide of glutathione and cystine (GSSSCy). 2. Four further fractions with activator activity were observed on ion-exchange chromatography of extracts prepared by methods similar to those described earlier [Neuberger et al. (1973)Biochem. J. 136,491-499]. These activators were generated by the extraction procedure. Two of them have been identified as trisulphanedisulphonate (S5O62-) and additional cystine trisulphide. 3. For the series of polysulphanedisulphonates (-O3S-Sn-SO3-, n greater than or equal to 1), activator activity at muM concentrations was exhibited only by compounds with n greater than 3. This, together with the results described above, indicates that for a compound R-Sn-R' (where R and R' are organic or inorganic groups) the only structural requirement for activity is n greater than or equal to 3. 4. Oxygenation of a semipanaerobic culture of R. spheroids for 1.5h before harvesting the cells produced a decrease of more than 90% in the cellular content of cystine trisulphide and glutathione trisulphides. 5. Chromatography on DEAE-Sephadex confirmed the presence of multiple forms of aminolaevulinate synthetase in extracts of R. spheroides [Tuboi et al. (1970) Arch. Biochem. Biophys. 138,147-154]. Oxygenation of a semi-anaerobic culture resulted in the disappearance of high-activity enzyme (a-form) and the accumulation of low-activity enzyme (b-form) in the cell. Spontaneous activation [Marriott et al. (1969) Biochem. J. 111,385-394] And activation by cystine trisulphide both resulted in the almost complete conversion of the b-form into the a-form.

scholarly journals Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. Binding of pyridoxal phosphate to 5-aminolaevulinate synthetase

1979 ◽  
Vol 177 (2) ◽  
pp. 661-671 ◽  
Author(s):  
R C Davies ◽  
A Neuberger
Keyword(s):  
Schiff Base ◽  
High Activity ◽  
Pyridoxal Phosphate ◽  
Enzymic Activity ◽  
Synthetase Activity ◽  
Dilute Solution ◽  
Concentrated Solutions ◽  
Active Centre ◽  
Rhodopseudomonas Spheroides ◽  
Low Activity

1. Pyridoxal 5′-phosphate is a cofactor essential for the enzymic activity of aminolaevulinate synthetase from Rhodopseudomonas spheroides. It also aids activation of the low-activity enzyme by trisulphides such as cystine trisulphide, whereas inactivation of enzyme is facilitated by its absence. 2. The fluorescence spectrum of purified high-activity enzyme is that expected for a pyridoxal phosphate–Schiff base, but the firmly bound cofactor does not appear to be at the active centre. In dilute solutions of enzyme this grouping is inaccessible to nucleophiles such as glycine, hydroxylamine, borohydride and cyanide, at pH 7.4. 3. An active-centre Schiff base is formed between enzyne and added pyridoxal phosphate, which is accessible to nucleophiles. Concentrated solutions of this enzyme–Schiff base on treatment with glycine yield apo- and semi-apoenzyme, which can re-bind pyridoxal phosphate. 4. Two types of binding of pyridoxal phosphate are distinguishable in dilute solution of enzyme, but these become indistinguishable when concentrated solutions are treated with cofactor. A change occurs in the susceptibility towards borohydride of the fluorescence of the “structural” pyridoxal phosphate. 5. One or two molecules of cofactor are bound per subunit of mol. wt. 50 000 in semiapo- or holo-enzyme. The fluorescence of pyridoxamine phosphate covalently bound to enzyme also indicates one to two nmol of reducible Schiff base per 7000 units of activity in purified and partially purified samples of enzyme. 6. Cyanide does not convert high-activity into low-activity enzyme, but with the enzyme-pyridoxal phosphate complex it forms a yellow fluorescent derivative that is enzymically active.

scholarly journals Metabolic interaction between amino acid deprivation and cisplatin synergistically reduces phosphoribosyl-pyrophosphate and augments cisplatin cytotoxicity

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Nisreen Wahwah ◽  
Debanjan Dhar ◽  
Hui Chen ◽  
Shunhui Zhuang ◽  
Adriano Chan ◽  
...  
Keyword(s):  
Amino Acid ◽  
Essential Amino Acid ◽  
De Novo ◽  
Synthetase Activity ◽  
Amino Acid Deprivation ◽  
Phosphoribosyl Pyrophosphate ◽  
Nucleotide Synthesis ◽  
Intracellular Phosphate ◽  
Dna Strand

AbstractCisplatin is a mainstay of cancer chemotherapy. It forms DNA adducts, thereby activating poly(ADP-ribose) polymerases (PARPs) to initiate DNA repair. The PARP substrate NAD+ is synthesized from 5-phosphoribose-1-pyrophosphate (PRPP), and we found that treating cells for 6 h with cisplatin reduced intracellular PRPP availability. The decrease in PRPP was likely from (1) increased PRPP consumption, because cisplatin increased protein PARylation and PARP1 shRNA knock-down returned PRPP towards normal, and (2) decreased intracellular phosphate, which down-regulated PRPP synthetase activity. Depriving cells of a single essential amino acid decreased PRPP synthetase activity with a half-life of ~ 8 h, and combining cisplatin and amino acid deprivation synergistically reduced intracellular PRPP. PRPP is a rate-limiting substrate for purine nucleotide synthesis, and cisplatin inhibited de novo purine synthesis and DNA synthesis, with amino acid deprivation augmenting cisplatin’s effects. Amino acid deprivation enhanced cisplatin’s cytotoxicity, increasing cellular apoptosis and DNA strand breaks in vitro, and intermittent deprivation of lysine combined with a sub-therapeutic dose of cisplatin inhibited growth of ectopic hepatomas in mice. Augmentation of cisplatin’s biochemical and cytotoxic effects by amino acid deprivation suggest that intermittent deprivation of an essential amino acid could allow dose reduction of cisplatin; this could reduce the drug’s side effects, and allow its use in cisplatin-resistant tumors.

Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides

1976 ◽  
Vol 273 (924) ◽  
pp. 79-98 ◽  
Keyword(s):  
High Activity ◽  
Photosynthetic Pigment ◽  
Synthetase Activity ◽  
Pigment Synthesis ◽  
Cellular Content ◽  
Rapid Fall ◽  
Activity Form ◽  
Oxygen Tensions ◽  
Rhodopseudomonas Spheroides ◽  
Low Activity

Rhodopseudomonas spheroides can grow in a defined medium with either light or oxygen as an energy source. Cells grown anaerobically or at very low oxygen tensions are rich in the photosynthetic pigment bacteriochlorophyll, whereas this pigment is virtually absent in cells grown under high oxygen tensions. Aminolaevulinate synthetase, the first enzyme on the pathway to bacteriochlorophyll, appears to play an important role in the control of bacteriochlorophyll synthesis. Thus, the enzyme has a high activity in extracts of pigmented cells and a low activity in extracts of non-pigmented cells. Further, oxygenation of a pigmented culture causes immediate cessation of pigment synthesis and produces a rapid fall in the activity of aminolaevulinate synthetase. This loss of activity appears to be due to the loss of an endogenous activator of the enzyme. Thus, pigmented cells contain cystine trisulphide, which at μM concentrations is an activator of aminolaevulinate synthetase, while oxygenation causes a rapid fall in the cellular content of this trisulphide. Cystathionase (EC 4 2.1.15) extracted from pigmented cells can catalyse the formation of cystine trisulphide from cystine, while rhodanese (EC 2.8.1.1) extracted from the same cells can catalyse the degradation of cystine trisulphide in the presence of sulphite to form cystine and thiosulphate. It is proposed that the cellular content of cystine trisulphide is controlled by changes in the levels of substrates for cystathionase and possibly rhodanese rather than changes in the amounts of these enzymes. Cystine trisulphide controls the activity of aminolaevulinate synthetase by converting a low-activity form of the enzyme (b-form) into a high-activity form (a-form). The fall in aminolaevulinate synthetase activity on oxygenation appears to be the result of cessation of conversion of b-form into a-form, along with a conversion of a-form into b-form. Factors affecting the equilibrium between the forms and the possible mechanisms for their interconversion are discussed.

Einfluß der Glucose auf die [14C] Thymidin-Einbaurate von Ehrlich-Ascitescarcinomzellen in vitro

1969 ◽  
Vol 08 (02) ◽  
pp. 196-206 ◽  
Author(s):  
Dieter. Kummer
Keyword(s):  
De Novo

ZusammenfassungIn nahezu glucosefreier Suspension von Ehrlich-Ascitescarcinomzellen bewirkt die Zufuhr von Glucose 2,5 × 10–4 bis 10–2 M:1. Hemmung der [14C] Thymidin-Einbaurate in die Zellen.2. Aktivierung des Ribonucleotid-Reductase-Systems und damit Stimulierung der Desoxyribonucleotidsynthese (auch der Thymidintriphosphat-de-novo-Synthese).3. Blockierung der Thymidinkinase über Endprodukthemmung, wodurch die Minderung des [14C] Thymidin-Einbaus in die Zellen erklärbar ist.

A Comparison of the in Vitro and in Vivo Thrombogenic Activity of Factor IX Concentrates Using Stasis (Wessler) and Non-Stasis Rabbit Models

1980 ◽  
Vol 44 (02) ◽  
pp. 081-086 ◽  
Author(s):  
C V Prowse ◽  
A E Williams
Keyword(s):  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Factor Ix ◽  
Coagulation System ◽  
In Vitro Tests ◽  
Clinical Use ◽  
Low Activity

SummaryThe thrombogenic effects of selected factor IX concentrates were evaluated in two rabbit models; the Wessler stasis model and a novel non-stasis model. Concentrates active in either the NAPTT or TGt50 in vitro tests of potential thrombogenicity, or both, caused thrombus formation in the Wessler technique and activation of the coagulation system in the non-stasis model. A concentrate with low activity in both in vitro tests did not have thrombogenic effects in vivo, at the chosen dose. Results in the non-stasis model suggested that the thrombogenic effects of factor IX concentrates may occur by at least two mechanisms. A concentrate prepared from platelet-rich plasma and a pyrogenic concentrate were also tested and found to have no thrombogenic effect in vivo.These studies justify the use of the NAPTT and TGt50 in vitro tests for the screening of factor IX concentrates prior to clinical use.

Особенности морфогенеза редкого вида Allium neriniflorum (Herb.) Backer в условиях in vitro

2019 ◽  
pp. 37-41 ◽  
Author(s):  
Альбина Шамсуновна Ахметова ◽  
Альфия Ануровна Зарипова
Keyword(s):  
De Novo

Показана возможность эффективного применения метода культуры тканей для размножения Allium neriniflorum (Herb.) Backer. Исследуемый вид является декоративным растением, размножение которого затруднено из-за низкой всхожести семян и ослабленной способности к формированию дочерних луковиц. Разработана технология клонального микроразмножения из стерильных луковиц. В качестве исходного материала использовали семена A. neriniflorum. Подобраны условия стерилизации, позволяющие достичь максимального числа (75 %) жизнеспособных эксплантов. Поверхностную стерилизацию проводили в ламинар-боксе с использованием в качестве стерилизующего агента 0,1 % раствор диацида. Семена сначала обрабатывали 70 % этанолом, затем стерилизующим раствором. Экспозиция стерилизующих растворов составляла от 5 до 9 мин. Показано, что способность к индуцированному морфогенезу существенно зависит от состава питательной среды. Максимальное число луковиц образовывалось на среде QL — 9 шт./эксплант. Исследуемые виды обладали высокой способностью к мультипликации и формированию полноценных растений при подобранных условиях культивирования in vitro. Выявленная морфогенетическая активность зачаточного побега, сегментов чешуй и донца стерильной луковицы A. neriniflorum, проявляющаяся в способности регенерировать побеги de novo, что возможно только в культуре in vitro, обеспечивает формирование полноценных луковиц. Луковицы, полученные in vitro, включали в последующие циклы микроразмножения. Культура тканей и органов in vitro позволяет размножать A. neriniflorum с более высоким коэффициентом размножения. От одной стерильной луковицы можно получить до 7000 луковиц в год. При традиционном вегетативном способе размножения материнская луковица формирует 1, редко 2 дочерние луковицы.

Study of Phosphocalcic Glasses from SiO2-CaO-P2O5 System with and Without Silver II. The bioactivity analysis by FTIR, SEM methods and microbiological study of silver-doped glasses

2017 ◽  
Vol 68 (6) ◽  
pp. 1188-1192
Author(s):  
Daniela Avram ◽  
Nicolae Angelescu ◽  
Dan Nicolae Ungureanu ◽  
Ionica Ionita ◽  
Iulian Bancuta ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Body Fluid ◽  
Pathogenic Bacteria ◽  
De Novo ◽  
Microbiological Examination ◽  
Doped Glasses ◽  
P2o5 System ◽  
Scanning Electron

The study in vitro of the glass powders bioactivity was performed by soaking them in simulated body fluid for 3 to 21 days at a temperature of 37�C and pH = 7.20. The synthesis de novo of hydroxyapatite, post soaking was confirmed by Fourier Transform Infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). The study of the antimicrobial activity was performed by microbiological examination on two strains of pathogenic bacteria involved in postoperative nosocomial infections.

Synthesis, Antimalarial Evaluation and SAR Study of Some 1,3,5-Trisubstituted Pyrazoline Derivatives

2019 ◽  
Vol 16 (10) ◽  
pp. 807-817 ◽  
Author(s):  
Shilpy Aggarwal ◽  
Deepika Paliwal ◽  
Dhirender Kaushik ◽  
Girish Kumar Gupta ◽  
Ajay Kumar
Keyword(s):  
Antimalarial Activity ◽  
Analysis Data ◽  
Docking Study ◽  
Maximum Activity ◽  
Elemental Analysis Data ◽  
Mass Spectral ◽  
Structure Activity ◽  
Pharmacokinetic Properties ◽  
Sar Study

The synthesis of a novel series of 1,3,5-trisubstitiuted pyrazoline was achieved by refluxing chalcone derivative with different heteroaryl hydrazines. The newly synthesized compounds were characterized by 1H NMR, 13CNMR, mass spectral and elemental analysis data. The synthetic series of novel pyrazoline hybrids was screened for in vitro schizont maturation assay against chloroquine sensitive 3D7 strain of Plasmodium falciparum. Most of the compounds showed promising in vitro antimalarial activity against CQ sensitive strain. The preliminary structure-activity relationship study showed that quinoline substituted analog at position N-1 showed maximum activity followed by benzothiazole substitution, while phenyl substitution lowers the antimalarial activity. The observed activity was persistent by the docking study on P. falciparum cystein protease falcipain-2. The pharmacokinetic properties were also studied using ADME prediction.

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