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Bio-statistical enhancement of acyl transfer activity of amidase for biotransformation of N-substituted aromatic amides
Abstract
Acyl transfer activity (ATA) of amidase transfers an acyl group of different amides to hydroxylamine to form the corresponding hydroxamic acid. Bacterial isolate BR-1 was isolated from cyanogenic plant Cirsium vulgare rhizosphere and identified as Pseudomonas putida BR-1 by 16S rDNA sequencing. This organism exhibited high ATA for the biotransformation of N-substituted aromatic amide to the corresponding hydroxamic acid. Optimization of media, tryptone (0.6%), inducer, pH 8.5, and a growth temperature 25 degrees C for 56 h, resulted in a 7-fold increase in ATA. Further, Response Surface Methodology (RSM) and multiple feeding approach (20 mM after 14 h) of inducer led to a 29% enhancement of ATA from this organism. The half life (t(1/2)) of this enzyme at 50 degrees C and 60 degrees C was 3 h and 1 h, respectively. The ATA of amidase of Pseudomonas putida BR-1 makes it a potential candidate for the production of a variety of N-substituted aromatic hydroxamic acid.
Alcaligenes sp. MTCC 10674 has been isolated as a nitrile degrading bacterium from the soil. Amidase of this organism exhibited dual activity i.e. hydrolase as well as acyl transfer. The acyl transfer activity of this organism has been used for the synthesis of variety of hydroxamic acids. Optimization of physiochemical parameters resulted into 30 folds increase in the acyl transfer activity of amidase (0.039 Umgdcm-1 to 1.17 Umgdcm-1). The acyl transfer activity of amidase of this organism has broader substrate affinity ranging from a variety of aliphatic amides (formamide, acetamide, propanamide etc.) to aromatic amides (benzamide, mandelamide, nicotinamide etc.) along with hydroxylamine for the biotransformation of these amides into corresponding hydroxamic acid. This enzyme is quite stable at 50°C with t1/2 for 8h and at 60°C this amidase have t1/2 for 1.30 h. The acyl transfer activity of amidase of Alcaligenes sp. MTCC 10674 under high temperature condition makes of potential application in developing a bioprocess for the production of variety of aliphatic and aromatic hydroxamic acid.
A yeast strain identified as Geotrichum sp. JR1 was able to use acetonitrile as the sole carbon and nitrogen source. The strain grew in 0.5 to 2M acetonitrile. Ammonia generation as enzymatic product during the strain growth indicates the presence of all acetonitrile degrading enzyme. Acetic acid and acetamide were detected during assays with the resting cells Cultivated in acetonitrile, indicating the presence of nitrile and amide degrading enzymes. This paper is the first to describe the use of acetonitrile as the sole carbon and nitrogen Source by a yeast.
In this work, acyltransferase activity of a new bacterial isolate Bacillus smithii strain IITR6b2 was utilized for the synthesis of nicotinic acid hydroxamate (NAH), a heterocyclic class of hydroxamic acid. NAH is an important pyridine derivative and has found its role as bioligand, urease inhibitor, antityrosinase, antioxidant, antimetastatic, and vasodilating agents. Amidase having acyltransferase activity with nicotinamide is suitable for nicotinic acid hydroxamate production. However, amidase can also simultaneously hydrolyze nicotinamide and nicotinic acid hydroxamate to nicotinic acid. Nicotinic acid is an undesirable by-product and thus any biocatalytic process involving amidase for nicotinic acid hydroxamate production needs to have high ratios of acyltransferase to amide hydrolase and acyltransferase to nicotinic acid hydroxamate hydrolase activity. Isolate Bacillus smithii strain IITR6b2 was found to have 28- and 12.3-fold higher acyltransferase to amide and hydroxamic acid hydrolase activities, respectively. This higher ratio resulted in a limited undesirable by-product, nicotinic acid (NA) synthesis. The optimal substrate/co-substrate ratio, pH, temperature, incubation time, and resting cells concentration were 200/250 mM, 7, 30 °C, 40 min, and 0.7 mgDCW ml(-1), respectively, and 94.5 % molar conversion of nicotinamide to nicotinic acid hydroxamate was achieved under these reaction conditions. To avoid substrate inhibition effect, a fed-batch process based on the optimized parameters with two feedings of substrates (200/200 mM) at 40-min intervals was developed and a molar conversion yield of 89.4 % with the productivity of 52.9 g h(-1) g DCW (-1) was achieved at laboratory scale. Finally, 6.4 g of powder containing 58.5 % (w/w) nicotinic acid hydroxamate was recovered after lyophilization and further purification resulted in 95 % pure product.
The bioprocess employing acyl transferase activity of intracellular amidase of BTP-5x MTCC 9225 was harnessed for the synthesis of pharmaceutically important acetohydroxamic acid. BTP-5x exhibited highest acyl transferase activity with acetamide: hydroxylamine in ratio of 1:5 in 0.1 M NaHPO/NaHPO buffer (pH 7.5) at 65°C. In one liter fed-batch reaction containing 1:5 ratio of two substrates total of eight feedings of 0.05 M/20 min of acetamide were made and it was found that maximum acetohydroxamic production was achieved at 3:5 ratios of substrate and cosubstrate. In 1 l bench scale batch reaction containing 0.3 M acetamide, 0.5 M hydroxylamine in 0.1 M NaHPO/NaHPO buffer (pH 7.5, 50°C, 400 rpm) and 0.5 mg/ml (dry cell weight) of whole cells of BTP-5x (as biocatalyst) resulted in an yield of 0.28 M of acetohydroxamic acid after 20 min reaction time at 50°C. The acetamide bioconversion rate was 90-95% (mol mol) and 51 g powder containing 40% (w/w) acetohydroxamic acid was recovered after lyophilization.
In this study, exopolysaccharides (EPSs) production was investigated by growing strains of Pseudomonas aeruginosa G1 and Pseudomonas putida G12 in medium containing various carbon sources such as glucose, mannose, fructose, and xylose. EPS production (192 and 182 mg/L, respectively) of these strains grown in PAP medium with 2% glycerol (v/v) was used as control. The highest EPS production of the two strains was found in the xylose containing medium. The effect of different concentrations [2–6% (w/v)] of xylose on EPS production of both strains was also studied. The maximum EPS yield (368 mg/l−1) of the strain G1 was recorded in 3% (w/v) xylose, while the highest yield EPS yield (262 mg/L−1) of the strain G12 was recorded in 2% (w/v) xylose. The monosaccharide compositions of EPS produced by the two strains were analyzed by HPLC. Strain G1 [2% (w/v) glycerol] was found to compose of neutral sugars (92.0%), acetylated amino sugars (8.0%), while strain G1 [3% (w/v) xylose] contained neutral sugars (99.2%), acetylated amino sugars (0.8%). However, the composition of strain G12 [2% (w/v) glycerol] was neutral sugars (96.8%), acetylated amino sugars (3.2%) and the strain G12 [2% (w/v) xylose] was found to contain neutral sugars (96.1%), acetylated amino sugars (3.9%).
Environmental pollution is growing more and more due to the indiscriminate and frequently deliberate release of hazardous, harmful substances. Research efforts have been devoted to develop new, low-cost, low-technology, eco-friendly treatments capable of reducing and even eliminating pollution in the atmosphere, the hydrosphere and soil environments. Among biological agents, enzymes have a great potentiality to effectively transform and detoxify polluting substances because they have been recognized to be able to transform pollutants at a detectable rate and are potentially suitable to restore polluted environments. This brief review will examine some classes of pollutants and enzymes capable of transforming them effectively into innocuous products. Particular attention will be devoted to pollutants with a high polluting potential such as polyphenols, nitriles, PAHs, cyanides and heavy metals. The enzymatic processes developed and implemented in some of these detoxification treatments will be examined in details. The main advantages as well as the main drawbacks that are still present in the extensive application of enzymes in the in situ restoration of polluted environments will be discussed.
Ninety three cultures of plant growth promoting rhizobacteria (PGPR) isolated from rhizosphere of Pisum sativum, among them one isolate identified as Pseudomonas putida was found to be potential amidase producer. The organism exhibited a battery of PGPR traits including enhanced production of plant growth hormone indoleacetic acid (IAA) and siderophore. P. putida MTCC 6809 exhibited both intracellular and extra-cellular amidase activity. The organism produced maximum extracellular amidase enzyme at 30°C and pH 7.5 in shaking state. The organism hydrolyzed a wide range of aliphatic amides that included acetamide, propionamide, acrylamide and butyramide. Acrylamide is a known carcinogen, teratogen and neurotoxicant and utilization of acrylamide by P. putida MTCC 6809 assume great importance. The organism is also tolerant to number of heavy metals at higher levels. These characteristics make P. putida MTCC 6809 an excellent candidate for field application in contaminated soil.
The opportunistic fungal pathogen Aspergillus fumigatus adapts to iron limitation by upregulation of iron uptake mechanisms including siderophore biosynthesis and downregulation of iron-consuming pathways to spare iron. These metabolic changes depend mainly on the transcription factor HapX. Consistent with the crucial role of iron in pathophysiology, genetic inactivation of either HapX or the siderophore system attenuates virulence of A. fumigatus in a murine model of aspergillosis. The differences in iron handling between mammals and fungi might serve to improve therapy and diagnosis of fungal infections.
The enantioselective amidase from Rhodococcus sp. strain R312 was produced in Escherichia coli and was purified in one chromatographic step. This enzyme was shown to catalyze the acyl transfer reaction to hydroxylamine from a wide range of amides. The optimum working pH values were 7 with neutral amides and 8 with alpha-aminoamides. The reaction occurred according to a Ping Pong Bi Bi mechanism. The kinetic constants demonstrated that the presence of a hydrophobic moiety in the carbon side chain considerably decreased the Km(amide) values (e.g., Km(amide) = 0.1 mM for butyramide, isobutyramide, valeramide, pivalamide, hexanoamide, and benzamide). Moreover, very high turnover numbers (kcat) were obtained with linear aliphatic amides (e.g., kcat = 333 s-1 with hexanoamide), whereas branched-side-chain-, aromatic cycle- or heterocycle-containing amides were sterically hindered. Carboxylic acids, alpha-amino acids, and methyl esters were not acyl donors or were very bad acyl donors. Only amides and hydroxamic acids, both of which contained amide bonds, were determined to be efficient acyl donors. On the other hand, the highest affinities of the acyl-enzyme complexes for hydroxylamine were obtained with short, polar or unsaturated amides as acyl donors (e.g., KmNH2OH = 20, 25, and 5 mM for acetyl-, alanyl-, and acryloyl-enzyme complexes, respectively). No acyl acceptors except water and hydroxylamine were found. Finally, the purified amidase was shown to be L-enantioselective towards alpha-hydroxy- and alpha-aminoamides.
Many plant-associated bacteria synthesize the phytohormone indoleacetic acid (IAA). While IAA produced by phytopathogenic
bacteria, mainly by the indoleacetamide pathway, has been implicated in the induction of plant tumors, it is not clear whether
IAA synthesized by beneficial bacteria, usually via the indolepyruvic acid pathway, is involved in plant growth promotion.
To determine whether bacterial IAA enhances root development in host plants, the ipdc gene that encodes indolepyruvate decarboxylase, a key enzyme in the indolepyruvic acid pathway, was isolated from the plant
growth-promoting bacterium Pseudomonas putida GR12-2 and an IAA-deficient mutant constructed by insertional mutagenesis. The canola seedling primary roots from seeds treated
with wild-type P. putida GR12-2 were on average 35 to 50% longer than the roots from seeds treated with the IAA-deficient mutant and the roots from
uninoculated seeds. In addition, exposing mung bean cuttings to high levels of IAA by soaking them in a suspension of the
wild-type strain stimulated the formation of many, very small, adventitious roots. Formation of fewer roots was stimulated
by treatment with the IAA-deficient mutant. These results suggest that bacterial IAA plays a major role in the development
of the host plant root system.
We revisit statistical tests for branches of evolutionary trees reconstructed upon molecular data. A new, fast, approximate likelihood-ratio test (aLRT) for branches is presented here as a competitive alternative to nonparametric bootstrap and Bayesian estimation of branch support. The aLRT is based on the idea of the conventional LRT, with the null hypothesis corresponding to the assumption that the inferred branch has length 0. We show that the LRT statistic is asymptotically distributed as a maximum of three random variables drawn from the chi(0)2 + chi(1)2 distribution. The new aLRT of interior branch uses this distribution for significance testing, but the test statistic is approximated in a slightly conservative but practical way as 2(l1- l2), i.e., double the difference between the maximum log-likelihood values corresponding to the best tree and the second best topological arrangement around the branch of interest. Such a test is fast because the log-likelihood value l2 is computed by optimizing only over the branch of interest and the four adjacent branches, whereas other parameters are fixed at their optimal values corresponding to the best ML tree. The performance of the new test was studied on simulated 4-, 12-, and 100-taxon data sets with sequences of different lengths. The aLRT is shown to be accurate, powerful, and robust to certain violations of model assumptions. The aLRT is implemented within the algorithm used by the recent fast maximum likelihood tree estimation program PHYML (Guindon and Gascuel, 2003).
An amidase (EC 3.5.1.4) in branch 2 of the nitrilase superfamily, from the thermophilic strain Geobacillus pallidus RAPc8, was produced at high expression levels (20 U/mg) in small-scale fermentations of Escherichia coli. The enzyme was purified to 90% homogeneity with specific activity of 1,800 U/mg in just two steps, namely, heat-treatment and gel permeation chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electron microscopic (EM) analysis of the homogenous enzyme showed the native enzyme to be a homohexamer of 38 kDa subunits. Analysis of the biochemical properties of the amidase showed that the optimal temperature and pH for activity were 50 and 7.0 degrees C, respectively. The amidase exhibited high thermal stability at 50 and 60 degrees C, with half-lives greater than 5 h at both temperatures. At 70 and 80 degrees C, the half-life values were 43 and 10 min, respectively. The amidase catalyzed the hydrolysis of low molecular weight aliphatic amides, with D: -selectivity towards lactamide. Inhibition studies showed activation/inhibition data consistent with the presence of a catalytically active thiol group. Acyl transfer reactions were demonstrated with acetamide, propionamide, isobutyramide, and acrylamide as substrates and hydroxylamine as the acyl acceptor; the highest reaction rate being with isobutyramide. Immobilization by entrapment in polyacrylamide gels, covalent binding on Eupergit C beads at 4 degrees C and on Amberlite-XAD57 resulted in low protein binding and low activity, but immobilization on Eupergit C beads at 25 degrees C with cross-linking resulted in high protein binding yield and high immobilized specific activity (80% of non-immobilized activity). Characterization of Eupergit C-immobilized preparations showed that the optimum reaction temperature was unchanged, the pH range was somewhat broadened, and stability was enhanced giving half-lives of 52 min at 70 degrees C and 30 min at 80 degrees C. The amidase has potential for application under high temperature conditions as a biocatalyst for D: -selective amide hydrolysis producing enantiomerically pure carboxylic acids and for production of novel amides by acyl transfer.
Using racemic tert-leucine amide as sole nitrogen source in minimal medium, 162 strains were isolated by enrichment techniques and shown to contain amidase activity. Among these isolates three D-amidase producers were found and identified as Variovorax paradoxus (two strains) and Klebsiella spec. The D-amidase from Variovorax paradoxus was purified to homogeneity by three chromatographic steps. With DL-Tle-amide as substrate Michaelis-Menten kinetics were observed with a K-M of 0.74 mM, a K-1 of 640 mM and a V-max of 1.4 U/mg. The amidase has a broad pH-optimum between 7 and 9.5 and a temperature optimum at 47 - 49 degreesC. The amidase hydrolyzed amino acid amides as well as carboxamides and 2-hydroxy acid amides. The stereoselectivity of the reaction was variable, however. Hydrolyzing DL-Tle-amide the enantiomeric ratio E was >200 resulting in D-Tle with an ee of >99% and up to 47% conversion. Similar results were obtained with DL-Leu-amide and DL-Val-amide while DL-Phe-amide was hydrolyzed with an enantiomeric ratio E of only 5.
Alcaligenes sp. MTCC 10674 has been isolated as a nitrile degrading bacterium from the soil. Amidase of this organism exhibited dual activity i.e. hydrolase as well as acyl transfer. The acyltransfer activity of this organism has been used for the synthesis of variety of hydroxamic acids. Optimization of physiochemical parameters resulted into 30 folds increase in the acyl transfer activity of amidase (0.039 Umgdcm-1 to 1.17 Umgdcm-1). The acyl transfer activity of amidase of this organism has broader substrate affinity ranging from a variety of aliphatic amides (formamide, acetamide, propanamide etc.) to aromatic amides (benzamide, mandelamide, nicotinamide etc.) along with hydroxylamine for the biotransformation of these amides into corresponding hydroxamic acid. This enzyme is quite stable at 50°C with t1/2 for 8h and at 60°C this amidase have t1/2 for 1.30 h. The acyl transfer activity of amidase of Alcaligenes sp. MTCC 10674 under high temperature condition makes of potential application in developing a bioprocess for the production of variety of aliphatic and aromatic hydroxamic acid.
The acyl transfer activity of the amidase of Alcaligenes sp. MTCC 10674 has been applied to the conversion of benzamide and hydroxylamine to benzohydroxamic acid. The unique features of the acyl transfer activity of this organism include its optimal activity at 50 °C and very high substrate (100 mM benzamide) and product (90 mM benzohydroxamic acid) tolerance among the hitherto reported enzymes. The bench scale production of benzohydroxamic acid was carried out in a fed-batch reaction (final volume 1 l) by adding 50 mM benzamide and 250 mM of hydroxylamine after every 20 min for 80 min in 0.1 M potassium phosphate buffer (pH 7.0) at 50 °C, using resting cells equal to 4.0 mg dcm/ml of reaction mixture. From 1 l of reaction mixture 33 g of benzohydroxamic acid was recovered with 24.6 g l(-1) h(-1) productivity. The acyl transfer activity of the amidase of Alcaligenes sp. MTCC 10674 and the process developed in the present study are of industrial significance for the enzyme-mediated production of benzohydroxamic acid.
Alcaligenes sp. MTCC 10674 was isolated as acetone cyanohydrin hydrolyzing bacterium from soil of orchid gardens of Himachal Pradesh. Acetone cyanohydrin hydrolyzing activity of this organism comprised nitrile hydratase and amidase activities. It exhibited higher substrate specificity towards aliphatic hydroxynitrile (acetone cyanohydrin) in comparison to arylaliphatic hydroxynitrile. Isobutyronitrile (40 mM) acted as a carbon source as well as inducer for growth of Alcaligenes sp. MTCC 10674 and expression of acetone cyanohydrin hydrolyzing activity. Optimization of culture condition using response surface methodology increased acetone cyanohydrin hydrolyzing activity by 1.3-fold, while inducer mediation approach increased the activity by 1.2-fold. The half life of this enzyme was 25 h at 15 °C. V
max and K
m value for acetone cyanohydrin hydrolyzing enzyme was 0.71 μmol mg−1 min−1 and 14.3 mM, when acetone cyanohydrin was used as substrate. Acetone cyanohydrin hydrolyzing enzyme encountered product inhibition and IC50 and K
i value were calculated to be 28 and 10.2 mM, respectively, when product α-hydroxyisobutyric acid was added in the reaction. Under optimized reaction conditions at 40 ml fed batch scale, 3 mg dcw ml−
resting cells of Alcaligenes sp. MTCC 10674 fully converted 0.33 M acetone cyanohydrin into α-hydroxyisobutyric acid (1.02 g) in 6 h 40 min. The characterization of acetone cyanohydrins hydrolyzing activity revealed that it comprises bienzymatic nitrile hydrolyzing system, i.e. nitrile hydratase and amidase for the production of α-hydroxyisobutyric acid from acetone cyanohydrin and maximum 70 % yield is being reported for the first time.
Plant roots serve a multitude of functions in the plant including anchorage, provision of nutrients and water, and production of exudates with growth regulatory properties. The root–soil interface, or rhizosphere, is the site of greatest activity within the soil matrix. Within this matrix, roots affect soil structure, aeration and biological activity as they are the major source of organic inputs into the rhizosphere, and are also responsible for depletion of large supplies of inorganic compounds. Roots are very complicated morphologically and physiologically, and their metabolites are often released in large quantities into the soil rhizosphere from living root hairs or fibrous root systems. Root exudates containing root-specific metabolites have critical ecological impacts on soil macro and microbiota as well as on the whole plant itself. Through the exudation of a wide variety of compounds, roots impact the soil microbial community in their immediate vicinity, influence resistance to pests, support beneficial symbioses, alter the chemical and physical properties of the soil, and inhibit the growth of competing plant species. In this review, we outline recent research on root exudation and the role of allelochemicals in the rhizosphere by studying the case of three plants that have been shown to produce allelopathic root exudates: black walnut, wheat and sorghum
A novel amidase gene, designated pamh, was cloned from Paracoccus sp. M-1. Site-directed mutagenesis and bioinformatic analysis showed that the PamH protein belonged to the amidase signature enzyme family. PamH was expressed in Escherichia coli, purified, and characterized. The molecular mass of PamH was determined to be 52 kDa with an isoelectric point of 5.13. PamH displayed its highest enzymatic activity at 45°C and at pH 8.0 and was stable within a pH range of 5.0–10.0. The PamH enzyme exhibited amidase activity, aryl acylamidase activity, and acyl transferase activity, allowing it to function across a very broad substrate spectrum. PamH was highly active on aromatic and short-chain aliphatic amides (benzamide and propionamide), moderately active on amino acid amides, and possessed weak urease activity. Of the anilides examined, only propanil was a good substrate for PamH. For propanil, the k
cat and K
m were 2.8 s−1 and 158 μM, respectively, and the catalytic efficiency value (k
cat/K
m) was 0.018 μM−1 s−1. In addition, PamH was able to catalyze the acyl transfer reaction to hydroxylamine for both amide and anilide substrates, including acetamide, propanil, and 4-nitroacetanilide; the highest reaction rate was shown with isobutyramide. These characteristics make PamH an excellent candidate for environmental remediation and an important enzyme for the biosynthesis of novel amides.
Enzymes are biologically produced proteinic substances having specific activation in which they combine with their substrates in such a stereoscopic position that they cause changes in the electronic configuration around certain susceptible bonds. Their significance in all spheres including soil, is worth tested and reported. In plant nutrition their role cannot be substituted by any other substance and its function is quite pragmatic in solubilizing and dissolving the much needed food in ionic forms for the very survival of animal and plant kingdom. World over, innumerable researchers have contributed their efforts in exploring enzymes. This paper reviews some of the important factors affecting its behaviour, reactions in soil environment, correlation with other enzymes and soil properties, preceded by its historical perspective and sources of production.
Isolation and characterization of a beta-lactamase (EC 3.5.2.6)-free, penicillin amidase (penicillin amidohydrolase, EC 3.5.1. 11)-producing organism is reported. The test strain was isolated by an enrichment technique with a substrate other than penicillins. The isolated strain belongs to the genus Alcaligenes. Phenylacetic acid was found to be the inducer of penicillin amidase. The amidase has a broad substrate spectrum. It is very active against penicillin G and semisynthetic cephalosporins, whereas penicillin V and semisynthetic penicillins acted moderately as a substrate. Immobilized cells of Alcaligenes sp. were shown to act as a reversible enzyme.
The use of some multiple-sequence alignments in phylogenetic analysis, particularly those that are not very well conserved, requires the elimination of poorly aligned positions and divergent regions, since they may not be homologous or may have been saturated by multiple substitutions. A computerized method that eliminates such positions and at the same time tries to minimize the loss of informative sites is presented here. The method is based on the selection of blocks of positions that fulfill a simple set of requirements with respect to the number of contiguous conserved positions, lack of gaps, and high conservation of flanking positions, making the final alignment more suitable for phylogenetic analysis. To illustrate the efficiency of this method, alignments of 10 mitochondrial proteins from several completely sequenced mitochondrial genomes belonging to diverse eukaryotes were used as examples. The percentages of removed positions were higher in the most divergent alignments. After removing divergent segments, the amino acid composition of the different sequences was more uniform, and pairwise distances became much smaller. Phylogenetic trees show that topologies can be different after removing conserved blocks, particularly when there are several poorly resolved nodes. Strong support was found for the grouping of animals and fungi but not for the position of more basal eukaryotes. The use of a computerized method such as the one presented here reduces to a certain extent the necessity of manually editing multiple alignments, makes the automation of phylogenetic analysis of large data sets feasible, and facilitates the reproduction of the final alignment by other researchers.
SUMMARY The synthesis of an inducible amidase by Pseudomonas mginosu 86021~ was studied in cultures growing exponentially in succinate medium. Induction by both the substrate inducer acetamide, and the non-substrate inducer N-acetylacetamide, was repressed by cyanoacetamide. Induction by 10-2M-N-acetylacetamide was significantly repressed by lO-%-cyano- acetamide, but repression of induction by 10-3~-acetamide required a tenfold excess of cyanoacetamide. Amidase synthesis in a medium in which acetamide was the sole carbon+nitrogen source was also repressed by cyanoacetamide, which under these conditions inhibited the growth of non-induced bacteria. Several tricarboxylic acid cycle intermediates, and related compounds, repressed amidase synthesis in exponentially growing organisms. Catabolite repression by propionate in succinate medium was decreased by increasing the concentration of acetamide. These findings are discussed in relation to general theories of regulation of microbial enzyme synthesis.
A fractional factorial design with eight trials was applied to select and model the effects of major factors, individually and in combination, on improving Tetrahymena thermophila growth and enzyme production. Regulated pH at 6.8 and olive oil at 0.5% (v/v) showed positive effects on fermentation, and increased cell growth parameters including generation time and maximal population formation. Lipase and protease production were also improved by these factors and were favoured by cultivation of Tetrahymena in darkness. This statistical experiment offers a beneficial and rapid screening procedure to select the most effective combination of factors influencing fermentation processes.
The role of root exuadates and allelochemicals in rhizosphere
- C Bertin
- X Yang
Bertin, C., Yang, X., and Weston, L. A. (2003) The role of root exuadates
and allelochemicals in rhizosphere. Plant and Soil, 256, 67–83.
Soil enzymes research: A review online
- A Zahir
- M Atteeq
Zahir, A., Atteeq, M., and Arshad, M. (2001) Soil enzymes research: A
review online. J. Biol. Sci., 1, 299-307.