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1.
The contents of adenine nucleotides, phosphagens and some glycolytic intermediates in resting muscles from vertebrates and invertebrates. 总被引:5,自引:0,他引:5
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The lowest contents of ATP and the lowest ATP/AMP concentration ratios are observed in the molluscan muscles that have very low rates of energy expenditure during contraction. The highest contents of ATP are observed in the extremely aerobic insect flight muscle and the extremely anaerobic pectoral muscle of the pheasant and domestic fowl. In general, the lowest ATP/AMP concentration ratios are observed for muscle in which the variation in the rate of energy utilization is small (e.g. some molluscan muscles, heart muscle); the highest ratios are observed in muscles in which this variation is large (lobster abdominal muscle, pheasant pectoral muscle, some insect flight muscles). This finding is consistent with the proposed role of AMP and the adenylate kinase reaction in the regulation of glycolysis. However, in the flight muscle of the honey-bee the ATP/AMP ratio is very low, so that glycolysis may be regulated by factors other than the variation in AMP concentration. The variation in the contents of arginine phosphate in muscle from the invertebrates is much larger than the variation in creatine phosphate in muscle from the vertebrates. The contents of hexose monophosphates and pyruvate are, in general, higher in the muscles of vertebrates than in those of the invertebrates. The contents of phosphoenolpyruvate are similar in all the muscles investigated, except for the honey-bee in which it is about 4-10-fold higher. The mass-action ratios for the reactions catalysed by phosphoglucoisomerase and adenylate kinase are very similar to the equilibrium constants for these reactions. Further, the variation in the mass-action ratios between muscles is small. It is concluded that these enzymes catalyse reactions close to equilibrium. However, the mass-action ratios for the reactions catalysed by phosphofructokinase and pyruvate kinase are much smaller than the equilibrium constants. The variation in the ratios between different muscles is large. It is concluded that these enzymes catalyse nonequilibrium reactions. Since the variation in the mass-action ratios for the reactions catalysed by the phosphagen kinases (i.e. creatine and arginine phosphokinases) is small, it is suggested that these reactions are close to equilibrium. 相似文献
2.
Peter Hedden 《Physiologia plantarum》1997,101(4):709-719
Gibberellins (GAs) are biosynthesised from the diterpene ent -kaurene by a series of oxidative reactions catalysed by two classes of enzymes. The early steps, involving transformations of highly hydrophobic substrates, are carried out by membrane-associated monooxygenases, probably involving cytochrome P450, whereas the later reactions are catalysed by soluble 2-oxoglutarate-dependent dioxygenases. Some reactions involving substrates, such as GA12 and GA12 -aldehyde, that have intermediate polarity are catalysed by enzymes in both classes. The monooxygenases and dioxygenases catalyse the same types of reactions: hydroxylation, desaturation, alcohol and aldehyde oxidation. For both enzyme classes, the oxidant is thought to be an oxyferryl species, depicted as Felv =O, that is derived from molecular oxygen by different mechanisms, the reducing power being supplied by NADPH in the case of cytochrome P450 monooxygenases and by the decarboxylation of 2-oxoglutarate to succinate for the dioxygenases. The recent availability of cDNA clones for several of the dioxygenases and the ability to prepare active enzymes by heterologous expression of cDNAs in Escherichia coli have provided new opportunities for examining the function of these enzymes. They have relatively low substrate specificity and, in many cases, are multifunctional. Consequently, fewer enzymes than expected are required to produce the large number of GA structures encountered in higher plants. In the present review, the major oxygenases of GA biosynthesis are described and their reactions are discussed in an attempt to rationalise this multifunctionality. 相似文献
3.
The aim of this work was to use preparations from germinating seeds of Pisum sativum to determine the apparent equilibrium constant of the reaction catalysed by sucrose-phosphate synthase (EC 2.4.1.14) and to compare this with the mass-action ratio of the reaction in the seeds. The apparent equilibrium constant ranged from 5.3 at 0.25 mM-MgCl2, pH 7.0, to 62 at 10 mM-MgCl2, pH 7.5. The sucrose phosphate content of the seeds, 23 nmol/g fresh wt., was determined by separating sucrose phosphate from sucrose by ion-exchange chromatography and then measuring the sucrose released by alkaline phosphatase. Comparison of equilibrium constants and mass-action ratios in the cotyledons of 38 h-germinated seeds showed that the reactions catalysed by glucose-6-phosphate isomerase, phosphoglucomutase and UDP-glucose pyrophosphorylase are close to equilibrium, and those catalysed by sucrose-phosphate synthase and sucrose phosphatase are considerably displaced from equilibrium in vivo. 相似文献
4.
Peter Geigenberger Silke Langenberger Ingo Wilke Dieter Heineke Hans W. Heldt Mark Stitt 《Planta》1993,190(4):446-453
Metabolites and enzyme activities were measured in the phloem sap exuding from a cut hypocotyl of germinating castor-bean (Ricinus communis L.) seedlings. The sap contained considerable quantities of adenine nucleotides, uridine nucleotides, uridine diphosphoglucose (UDPGlc), all the major phosphorylated metabolites required for glycolysis, fructose-2,6-bisphosphate and pyrophosphate. Supplying 200 mM glucose instead of sucrose to the cotyledons resulted in high concentrations of glucose in the sap, but did not modify the metabolite levels. In contrast, when 200 mM fructose was supplied we found only a low level of fructose but a raised sucrose concentration in the sap, which was accompanied by a three-to fourfold decrease of UDPGlc, and an increase of pyrophosphate, UDP and UTP. The measured levels of metabolites are used to estimate the molar mass action ratios and in-vivo free-energy change associated with the various reactions of sucrose breakdown and glycolysis in the phloem. It is concluded that the reactions catalysed by ATP-dependent phosphofructokinase and pyruvate kinase are removed from equilibrium in the phloem, whereas the reactions catalysed by sucrose synthase, UDPGlc-pyrophosphorylase, phosphoglucose mutase, phosphoglucose isomerase, aldolase, triose-phosphate isomerase, phosphoglycerate mutase and enolase are close to equilibrium within the conducting elements of the phloem. Since the exuded sap contained negligible or undetectable activities of the enzymes, it is concluded, that the responsible proteins are bound, or sequesterd behind plasmodesmata, possibly in the companion cells. It is argued that sucrose mobilisation via a reversible reaction catalysed by sucrose synthase is particularily well suited to allow the rate of sucrose breakdown in the phloem to respond to changes in the metabolic requirement for ATP, and for UDPGlc during callose production. It is also calculated that the transport of nucleotides in the phloem sap implies that there must be a very considerable uptake or de-novo biosynthesis of these cofactors in the phloem. 相似文献
5.
J. Tramper 《Trends in biotechnology》1985,3(2):45-50
Many interesting chemical reactions that are difficult or impossible to establish by ordinary chemical means are efficiently catalysed by enzymes in microbial, plant and animal, and human cells. Why so few enzymes are used as catalysts in syntheses and whether this is likely to change in the future, is discussed here. 相似文献
6.
7.
The aim of this work was to identify the regulatory reactions of glycolysis in potato tubers. The amounts of glycolytic intermediates in aerobic and anoxic tubers were measured in freeze-clamped samples of tissue. Comparison of mass—action ratios with apparent equilibrium constants showed that in vivo the reactions catalysed by glucosephosphate isomerase, phosphoglycerate mutase and enolase were close to equilibrium. The ratios fructose-1,6-bisphosphate:fructose 6-phosphate, and pyruvate:phosphoenolpyruvate, respectively, showed that the reactions catalysed by phosphofructokinase and pyruvate kinase were considerably displaced from equilibrium. Stimulation of glycolysis by placing tubers in an atmosphere of nitrogen led to significant declines in their contents of fructose-6-phosphate and phosphoenolpyruvate. It is concluded that phosphofructokinase plays a dominant role in regulating entry into glycolysis, and that pyruvate kinase may regulate exit from glycolysis and the oxidative pentose phosphate pathway. Cold-induced sweetening of the tubers is discussed in the light of the above conclusions. 相似文献
8.
The mechanisms of reductive carboxylation reactions. Carbon dioxide or bicarbonate as substrate of nicotinamide-adenine dinucleotide phosphate-linked isocitrate dehydrogenase and `malic'' enzyme 总被引:5,自引:3,他引:2
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1. A simple kinetic method was devised to show whether dissolved CO(2) or HCO(3)- ion is the substrate in enzyme-catalysed carboxylation reactions. 2. The time-course of the reductive carboxylation of 2-oxoglutarate by NADPH, catalysed by isocitrate dehydrogenase, was studied by a sensitive fluorimetric method at pH7.3 and pH6.4, with large concentrations of substrate and coenzyme and small carbon dioxide concentrations. 3. Reaction was initiated by the addition of carbon dioxide in one of three forms: (i) as the dissolved gas in equilibrium with bicarbonate; (ii) as unbuffered bicarbonate solution; (iii) as the gas or as an unbuffered solution of the gas in water. Different progress curves were obtained in the three cases. 4. The results show that dissolved CO(2) is the primary substrate of the enzyme, and that HCO(3)- ion is at best a very poor substrate. The progress curves are in quantitative agreement with this conclusion and with the known rates of the reversible hydration of CO(2) under the conditions of the experiments. The effects of carbonic anhydrase confirm the conclusions. 5. Similar experiments on the reductive carboxylation of pyruvate catalysed by the ;malic' enzyme show that dissolved CO(2) is the primary substrate of this enzyme also. 6. The results are discussed in relation to the mechanisms of these enzymes, and the effects of pH on the reactions. 7. The advantages of the method and its possible applications to other enzymes involved in carbon dioxide metabolism are discussed. 相似文献
9.
10.
Eugne H.J.M. Jansen P. de Fluiter 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1992,580(1-2):325-346
The reactions catalysed by the various cytochrome P-450 enzymes are reviewed with respect to the analysis of products by high-performance liquid chromatography (HPLC). Especially biotransformation reactions of purified cytochrome P-450 enzymes in a reconstituted system and in microsomes mainly of rat liver origin are considered. Emphasis is put on the specificity of product formation due to the individual isozymes of cytochrome P-450. It is shown that the presence of eight cytochrome P-450 isozymes can be monitored and determined by specific product formation after HPLC analysis, which is an important parameter in toxicological studies. 相似文献
11.
Benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus. Purification and preliminary characterization. 总被引:2,自引:2,他引:0
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A quick, reliable, purification procedure was developed for purifying both benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from a single batch of Acinetobacter calcoaceticus N.C.I.B. 8250. The procedure involved disruption of the bacteria in the French pressure cell and preparation of a high-speed supernatant, followed by chromatography on DEAE-Sephacel, affinity chromatography on Blue Sepharose CL-6B and Matrex Gel Red A, and finally gel filtration through a Superose 12 fast-protein-liquid-chromatography column. The enzymes co-purified as far as the Blue Sepharose CL-6B step were separated on the Matrex Gel Red A column. The final preparations of benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II gave single bands on electrophoresis under non-denaturing conditions or on SDS/polyacrylamide-gel electrophoresis. The enzymes are tetramers, as judged by comparison of their subunit (benzyl alcohol dehydrogenase, 39,700; benzaldehyde dehydrogenase II, 55,000) and native (benzyl alcohol dehydrogenase, 155,000; benzaldehyde dehydrogenase II, 222,500) Mr values, estimated by SDS/polyacrylamide-gel electrophoresis and gel filtration respectively. The optimum pH values for the oxidation reactions were 9.2 for benzyl alcohol dehydrogenase and 9.5 for benzaldehyde dehydrogenase II. The pH optimum for the reduction reaction for benzyl alcohol dehydrogenase was 8.9. The equilibrium constant for oxidation of benzyl alcohol to benzaldehyde by benzyl alcohol dehydrogenase was determined to be 3.08 x 10(-11) M; the ready reversibility of the reaction catalysed by benzyl alcohol dehydrogenase necessitated the development of an assay procedure in which hydrazine was used to trap the benzaldehyde formed by the NAD+-dependent oxidation of benzyl alcohol. The oxidation reaction catalysed by benzaldehyde dehydrogenase II was essentially irreversible. The maximum velocities for the oxidation reactions catalysed by benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II were 231 and 76 mumol/min per mg of protein respectively; the maximum velocity of the reduction reaction of benzyl alcohol dehydrogenase was 366 mumol/min per mg of protein. The pI values were 5.0 for benzyl alcohol dehydrogenase and 4.6 for benzaldehyde dehydrogenase II. Neither enzyme activity was affected when assayed in the presence of a range of salts. Absorption spectra of the two enzymes showed no evidence that they contain any cofactors such as cytochrome, flavin, or pyrroloquinoline quinone. The kinetic coefficients of the purified enzymes with benzyl alcohol, benzaldehyde, NAD+ and NADH are also presented. 相似文献
12.
Rate-limiting steps in the cytidine pathway for the synthesis of phosphatidylcholine and phosphatidylethanolamine. 总被引:6,自引:3,他引:3
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J P Infante 《The Biochemical journal》1977,167(3):847-849
An analysis of the available data on the cytidine pathway for the synthesis of phosphatidylcholine and phosphatidylethanolamine, by the logic derived from the theoretical principles of metabolic regulation, shows that the first two reactions catalysed by choline (ethanolamine) kinase and phosphocholine (phosphoethanolamine) cytidylyltransferase are rate-limiting, whereas the phosphocholine (phosphoethanolamine) transferase step is near equilibrium in rat liver. 相似文献
13.
Progesterone and the metabolic control of the lactose biosynthetic pathway during lactogenesis in the rat 总被引:11,自引:8,他引:3
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1. Lactogenesis was initiated in pregnant rats by ovariectomy, thereby causing progesterone withdrawal, after which the mammary tissue was analysed for contents of enzymes and metabolites concerned with the biosynthesis of lactose. 2. Lactose synthesis increased about 126-fold with little or no accompanying change in the contents of most metabolic intermediates or in the adenine nucleotide energy charge. 3. Comparison of mass-action ratios with equilibrium constants showed that phosphoglucomutase (EC 2.7.5.1), UDP-glucose pyrophosphorylase (EC 2.7.7.9) and UDP-glucose epimerase (EC 5.1.3.2.) catalysed reactions close to equilibrium. Nucleoside diphosphokinase (EC 2.7.4.6.) activity was very high and probably equilibrates the UTP-UDP and ATP-ADP couples. Lactose synthetase and hexokinase (EC 2.7.1.1) appeared to catalyse rate-limiting reactions. 4. Large increases were seen of UDP-glucose pyrophosphorylase (5-fold), lactose synthetase A protein (3.8-fold) and alpha-lactalbumin (28-fold), but not of hexokinase, phosphoglucomutase, UDP-glucose epimerase, nucleoside diphosphokinase or glucose 6-phosphate dehydrogenase (EC 1.1.1.49) activities. 5. It appeared that the increased lactose synthesis was largely accounted for by the increased lactose synthetase A protein activity and alpha-lactalbumin. 相似文献
14.
Transient-time analysis of substrate-channelling in interacting enzyme systems. 总被引:1,自引:0,他引:1
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The kinetics of dynamically interacting enzyme systems is examined, in the light of increasing evidence attesting to the widespread occurrence of this mode of organization in vivo. The transient time, a key phenomenological parameter for the coupled reaction, is expressed as a function of the lifetime of the intermediate substrate. The relationships between the transient time and the pseudo-first-order rate constants for the coupled reaction by the complexed and uncomplexed enzyme species are indicative of the mechanism of intermediate transfer ('channelling'). In a dynamically interacting enzyme system these kinetic parameters are composite functions of those for the processes catalysed by the complex and by the separated enzymes. The mathematical paradigm can be extended to a linear sequence of N coupled reactions catalysed by dynamically (pair-wise) interacting enzymes. 相似文献
15.
Ulla Christensen 《Biocatalysis and Biotransformation》1993,8(3):229-238
The steady state rate equations of transfer reactions catalysed by enzymes that follow the serine proteinase reaction mechanism in their hydrolysis reactions, have been solved and integrated. The integrated equations allow for calculations of maximal yields of product and of the time, tmax, at which that yield is present in a given reaction mixture. These important quantities have not been dealt with in previous theoretical studies of such systems. 相似文献
16.
The flux of serine biosynthesis in the liver of the normal rabbit, and of the rat on a low protein diet, is most sensitive to the activity of phosphoserine phosphatase (flux control coefficient up to 0.97), the last of the three enzymes in the pathway after it branches from glycolysis. The concentration of the pathway product, serine, has a strong controlling influence on the flux (response coefficient up to -0.64) through feedback inhibition at this step. The pathway is therefore controlled primarily by the demand for serine rather than the supply of the pathway precursor, 3-phosphoglycerate. Under conditions where there is a lower biosynthetic flux, the flux control coefficients of the first two enzymes of the pathway are increased, and are probably dominant in the rat on a normal diet. In rabbit liver, when ethanol is used to inhibit serine biosynthesis, control can be distributed between the three enzymes, even though the reactions catalysed by the first two remain close to equilibrium. Apart from their intrinsic value in aiding the understanding of the regulation of mammalian serine metabolism, our findings illustrate the danger of assuming that there are invariant design principles in the regulation of metabolic pathways, such as feedback control on the first step after a branch. 相似文献
17.
The formation of a carbon-carbon bond is an essential step in the biosynthetic pathways by which fatty acids and polyketides are made. The thiolase superfamily enzymes catalyse this carbon-carbon-bond formation via a thioester-dependent Claisen-condensation-reaction mechanism. In this way, fatty-acid chains and polyketides are made by sequentially adding simple building blocks, such as acetate units, to the growing molecule. A common feature of these enzymes is a reactive cysteine residue that is transiently acylated in the catalytic cycle. The wide catalytic diversity of the thiolase superfamily enzymes is of great interest. In particular, the type-III polyketide synthases make complicated compounds of great biological importance using multiple, subsequent condensation reactions, which are all catalysed in the same active-site cavity. The crucial metabolic importance of the bacterial fatty-acid-synthesizing enzymes stimulates in-depth studies that aim to develop efficient anti-bacterial drugs. 相似文献
18.
The following processes are discussed in this article: enzyme-catalysed hydrolyses of carboxylic acid esters and amides, phosphate esters, nitriles and epoxides; esterification and inter-esterification reactions catalysed by enzymes; reduction of ketones to secondary alcohols using whole-cell systems or isolated dehydrogenases; oxidation of alicyclic and aromatic substrates using mono-oxygenases and dioxygenases in bacteria and fungi including enzyme-catalysed Baeyer-Villiger oxidations; aldol reactions, formation of optically active cyanohydrins and enzyme-catalysed acyloin type reactions. The use of these biocatalytic methods for the stereo-controlled preparation of important target structures is reviewed and some of the future directions for the biotransformation area are discussed. 相似文献
19.
Handan Melike D?nerta? Sergio Martínez Cuesta Syed Asad Rahman Janet M. Thornton 《PloS one》2016,11(2)
The relationship between enzyme-catalysed reactions and the Enzyme Commission (EC) number, the widely accepted classification scheme used to characterise enzyme activity, is complex and with the rapid increase in our knowledge of the reactions catalysed by enzymes needs revisiting. We present a manual and computational analysis to investigate this complexity and found that almost one-third of all known EC numbers are linked to more than one reaction in the secondary reaction databases (e.g., KEGG). Although this complexity is often resolved by defining generic, alternative and partial reactions, we have also found individual EC numbers with more than one reaction catalysing different types of bond changes. This analysis adds a new dimension to our understanding of enzyme function and might be useful for the accurate annotation of the function of enzymes and to study the changes in enzyme function during evolution. 相似文献
20.
S N Kochetkov A R Abduragimov A G Gabibov E S Severin 《Molekuliarnaia biologiia》1985,19(6):1562-1568
The catalytic subunit of cAMP-dependent protein kinase from pig brain was shown to catalyse an isotope exchange reaction ATP in equilibrium with ADP. The kinetic parameters of the exchange were determined. The enzyme can also use GTP as the donor substrate; phosphotransferase and "GTPase" reactions were investigated. Based on the kinetic data obtained in this and in the previous paper the free energy profiles of protein kinase catalysed reactions are discussed. 相似文献