Microbial metabolism of phenolic amines: degradation of dl-synephrine by an unidentified arthrobacter

Microorganisms capable of degrading dl-synephrine were isolated from soil of Citrus gardens by enrichment culture, with dl-synephrine as the sole source of carbon and nitrogen. An organism which appears to be an arthrobacter, but which cannot be identified with any of the presently recognized species was predominant in these isolates. It was found to metabolize synephrine by a pathway involving p-hydroxyphenylacetaldehyde, p-hydroxyphenylacetic acid, and 3,4-dihydroxyphenylacetic acid as intermediates. Some of the enzymes of this pathway were demonstrated in cell-free extracts. An aromatic oxygenase, which could also be readily obtained in a cell-free system, was found to degrade 3,4-dihydroxyphenylacetic acid by meta cleavage.

Microbial transformations of acyclic monoterpenes

Microbial degradation of geraniol, citronellol, linalool and their corresponding acetates, structurally modified linalool and linalyl acetate, α-terpineol and β-myrcene are presented. Oxygenative and prototropic rearrangements are normally observed during the microbial metabolism of monoterpenes. Three types of oxygenation reactions are observed, namely, (a) allylic oxygenation (b) oxygenation on a double bond and (c) addition of water across the double bond. The studies indicate commonality in the reaction types or processes occurring during the metabolism of various related monoterpenes and also establish the convergence of degradative pathways at a central catabolic intermediate.

Cells Producing Their Own Nemesis: Understanding Methylglyoxal Metabolism

Methylglyoxal, which is technically known as 2-oxopropanal or pyruvaldehyde, shows typical reactions of carbonyl compounds as it has both an aldehyde and a ketone functional group. It is an extremely cytotoxic physiological metabolite, which is generated by both enzymatic and nonenzymatic reactions. The deleterious nature of the compound is due to its ability to glycate and crosslink macromolecules like protein and DNA, respectively. However, despite having toxic effects on cellular processes, methylglyoxal retains its efficacy as an anticancer drug. Indeed, methylglyoxal is one of the well-known anticancer therapeutic agents used in the treatment. Several studies on methylglyoxal biology revolve around the manifestations of its inhibitory effects and toxicity in microbial growth and diabetic complications, respectively. Here, we have revisited the chronology of methylglyoxal research with emphasis on metabolism of methylglyoxal and implications of methylglyoxal production or detoxification on bacterial pathogenesis and disease progression. (C) 2014 IUBMB Life, 66(10): 667-678, 2014

Structural and functional analysis of two universal stress proteins YdaA and YnaF from Salmonella typhimurium: possible roles in microbial stress tolerance

In many organisms ``Universal Stress Proteins'' CUSPS) are induced in response to a variety of environmental stresses. Here we report the structures of two USPs, YnaF and YdaA from Salmonella typhimurium determined at 1.8 angstrom and 2.4 angstrom resolutions, respectively. YnaF consists of a single USP domain and forms a tetrameric organization stabilized by interactions mediated through chloride ions. YdaA is a larger protein consisting of two tandem USP domains. Two protomers of YdaA associate to form a structure similar to the YnaF tetramer. YdaA showed ATPase activity and an ATP binding motif G-2X-G-9X-G(S/T/N) was found in its C-terminal domain. The residues corresponding to this motif were not conserved in YnaF although YnaF could bind ATP. However, unlike YdaA, YnaF did not hydrolyse ATP in vitro. Disruption of interactions mediated through chloride ions by selected mutations converted YnaF into an ATPase. Residues that might be important for ATP hydrolysis could be identified by comparing the active sites of native and mutant structures. Only the C-terminal domain of YdaA appears to be involved in ATP hydrolysis. The structurally similar N-terminal domain was found to bind a zinc ion near the segment equivalent to the phosphate binding loop of the C-terminal domain. Mass spectrometric analysis showed that YdaA might bind a ligand of approximate molecular weight 800 daltons. Structural comparisons suggest that the ligand, probably related to an intermediate in lipid A biosynthesis, might bind at a site close to the zinc ion. Therefore, the N-terminal domain of YdaA binds zinc and might play a role in lipid metabolism. Thus, USPs appear to perform several distinct functions such as ATP hydrolysis, altering membrane properties and chloride sensing. (C) 2015 Elsevier Inc. All rights reserved.

UVB irradiation alters the activities and kinetic properties of the enzymes of energy metabolism in rat lens during aging

Ultraviolet (UV) radiation is one of the major risk factors of cataract (loss of eye-lens transparency). The influence of UVB radiation (300 nm, 100 mu W cm(-2)) on the activity and apparent kinetic constants (K-m and V-max) of rat lens hexokinase (HK;EC2.7.1.1), phosphofructokinase (PFK;EC2.7.1.11), isocitrate dehydrogenase (ICDH;EC1.1.1.41) and malate dehydrogenase (MDH;EC1.1.1.37) of energy metabolism has been investigated by irradiating the lens homogenate of three-and 12-month-old rats. In the three-month-old group specific activities of HK and PFK are reduced by 56 and 43 %, respectively, and there is no change in ICDH and MDH activities after a 24 h exposure. On the other hand, in the 12-month-old group the decreases are 72, 71, 24 and 16 % for HK, PFK. ICDH and MDH, respectively. UVB irradiation increases the apparent K-m of HK and PFK (in both age groups), whereas the K-m of ICDH and MDH is not altered. While the decrease in V-max of these enzymes due to UVB exposure is only marginal in three-month-old rats, it is more pronounced (significant) in 12-month-old rats. A similar decrease in enzyme activities of HK and PFK is also observe upon UVB exposure of the intact rat lens. The photoinduced changes in energy metabolism may in turn have a bearing on lens transparency, particularly at an older age.

Expression Profiling of Nucleotide Metabolism-Related Genes in Human Breast Cancer Cells After Treatment with 5-Fluorouracil

5-Fluorouracil (5-FU) is one of the most widely used drugs for treatment of cancers, including breast cancer that exhibits its anticancer activity by inhibiting DNA synthesis and also incorporated into DNA and RNA. The objective of this investigation was to find out the total nucleotide metabolism genes regulated by 5-FU in breast cancer cell line. The breast cancer cell line MCF-7 was treated with the drug 5-FU. To analyze the expression of genes, we have conducted the experiment using 1.7k and 19k human microarray slide and confirmed the expression of genes by semiquantitative reverse transcription-polymerase chain reaction. The expression of 44 genes involved in the nucleotide metabolism pathway was quantified. Of these 44 genes analyzed, transcription of 6 genes were upregulated and 9 genes were downregulated. Earlier studies revealed that the transcription of genes for key enzymes like thymidylate synthase, thymidinekinase, and dihydropyrimidine dehydrogenase are regulated by 5-FU. This study identified some novel genes like thioredoxin reductase, ectonucleotide triphosphate dephosphorylase, and CTP synthase are regulated by 5-FU. The data also reveal large-scale perturbation in transcription of genes not involved directly in the known mechanism of action of 5-FU.

Metabolism of 1,8-Cineole in Rat: Its Effects on Liver and Lung Microsomal Cytochrome P-450 Systems

The monoterpene cyclic ether, cineole (l,8-cineole, I) also known as eucalyptol, is a component of many essential oils and is widely distributed in nature. It is extensively used in pharmaceutical preparations for external application and also as a nasal spray. It was reported earlier that cineole when administered to sheep may be largely oxidized in the system (Scheline 1978). However the mode of metabolism of cineole is not known. Hence the present study was undertaken to investigate the metabolic fate of this ubiquitous terpenoid following its administration to rats by gastric intubation.

Comparative phytotoxicity of nitrophenolic soil pollutants and their microbial metabolites to the growth of cucumber

2,4-Dinitrophenol and paranitrophenol are two major soil pollutants which are known to be metabolized by different soil microbes. Relative phytotoxicities of these parent compounds and their metabolic transformation products to the growth of cucumber seedlings were assessed. It was evident that such microbial transformations widely occurring in the soil are effective detoxification reactions and are beneficial for the plants.

Microbial transformation of a fungicide-derived amine, dimethyl-p-phenylene diamine by Alcaligenes DM4

Abstract Microbial transformation of N, N-dimethyl-p-phenylene diamine (DMPDA), a microbial product formed from the fungicide fenaminosulf (p-dimethylaminobenzenediazo sodium sulfonate) was studied by enriching microbes in soils treated with the amine. Microorganisms isolated from DMPDA-treated soil belonged to the genera of Micrococcus, Alcaligenes, and Corynebacterium. Of the various isolates, Alcaligenes DM4 showed maximal growth on DMPDA utilizing it as sources of carbon and nitrogen. When grown in mineral salts basal medium containing 0.05% DMPDA to serve as carbon and nitrogen sources, Alcaligenes DM4 grew exponentially up to 18 h. Even though the characterization of the complete pathway of microbial degradation of DMPDA could not be carried out due to the auto-oxidation of the compound, the initial transformation product of DMPDA by Alcaligenes DM4 has been identified as a dimer. The dimer is generated into the culture medium presumably by the extra-cellular oxidase of Alcaligenes DM4. It is suggested that the risk-benefit evaluation on the use of fenaminosulf is to be made taking into consideration the microbial transformations of the fungicide.

Microbial susceptibility of condensation products of carboxy-terminated polybutadiene and glycerol

Keeping in view the prospects of biodegradable polymers, a polymer was synthesized by the condensation of carboxy-terminated polybutadiene (CTPB) of Mnsim-5000 with glycerol and tested for its microbial susceptibility. The results of end group estimations and viscosity measurements indicated a quantitative reaction between the two reactants under experimental conditions. The clear-zone method was employed in this investigation to test biodegradability. Two strains of Serratia and three strains of Staphylococcus did show a clear zone surrounding the colony. However, the microbial growth was found to diminish after 4 or 5 days.

Partial purification and properties of a transamidinase from Lathyrus sativus seedlings. Involvement in homoarginine metabolism and amine interconversions

A transamidinase was purified 463-fold from Lathyrus sativus seedlings by affinity chromatography on homoarginine--Sepharose. The enzyme exhibited a wide substrate specificity, and catalysed the reversible transfer of the amidino groups from donors such as arginine, homoarginine and canavanine to acceptors such as lysine, putrescine, agmatine, cadaverine and hydroxylamine. The enzyme could not be detected in the seeds, and attained the highest specific activity in the embryo axis on day 10 after seed germination. Its thiol nature was established by strong inhibition by several thiol blockers and thiol compounds in the presence of ferricyanide. In the absence of an exogenous acceptor, it exhibited weak hydrolytic activity towards arginine. It had apparent mol.wt. 210000, and exhibited Michaelis--Menten kinetics with Km 3.0 mM for arginine. Ornithine competitively inhibited the enzyme, with Ki 1.0 mM in the arginine--hydroxylamine amidino-transfer reaction. Conversion experiments with labelled compounds suggest that the enzyme is involved in homoarginine catabolism during the development of plant embryo to give rise to important amino acids and amine metabolites. Presumptive evidence is also provided for its involvement in the biosynthesis of the guanidino amino acid during seed development. The natural occurrence of arcain in L. sativus and mediation of its synthesis in vitro from agmatine by the transamidinase are demonstrated.

Linalyl Acetate Is Metabolized by Pseudomonas incognita with the Acetoxy Group Intact

Metabolism of linalyl acetate by Pseudomonas incognita isolated by enrichment culture on the acyclic monoterpene alcohol linalool was studied. Biodegradation of linalyl acetate by this strain resulted in the formation of linalool, linalool- 8-carboxylic acid, oleuropeic acid, and A5-4-acetoxy-4-methyl hexenoic acid. Cells adapted to linalyl acetate metabolized linalyl acetate-8-aldehyde to linalool- 8-carboxylic acid, linalyl acetate-8-carboxylic acid, A5-4-acetoxy-4-methyl hexenoic acid, and geraniol-8-carboxylic acid. Resting cell suspensions previously grown with linalyl acetate oxidized linalyl acetate-8-aldehyde to linalyl acetate-8- carboxylic acid, A5-4-acetoxy-4-methyl hexenoic acid, and pyruvic acid. The crude cell-free extract (10,000 g of supernatant), obtained from the sonicate of linalyl acetate-grown cells, was shown to contain enzyme systems responsible for the formation of linalyl acetate-8-carboxylic acid and linalool-8-carboxylic acid from linalyl acetate. The same supernatant contained NAD-linked alcohol and aldehyde dehydrogenases involved in the formation of linalyl acetate-8-aldehyde and linalyl acetate-8-carboxylic acid, respectively. On the basis of various metabolites isolated from the culture medium, resting cell experiments, growth and manometric studies carried out with the isolated metabolites as well as related synthetic analogs, and the preliminary enzymatic studies performed with the cellfree extract, a probable pathway for the microbial degradation of linalyl acetate with the acetoxy group intact is suggested.

Lipids of nervous tissue: composition and metabolism

As indicated in the Introduction, the many significant developments in the recent past in our knowledge of the lipids of the nervous system have been collated in this article. That there is a sustained interest in this field is evident from the rather long bibliography which is itself selective. Obviously, it is not possible to summarize a review in which the chemistry, distribution and metabolism of a great variety of lipids have been discussed. However, from the progress of research, some general conclusions may be drawn. The period of discovery of new lipids in the nervous system appears to be over. All the major lipid components have been discovered and a great deal is now known about their structure and metabolism. Analytical data on the lipid composition of the CNS are available for a number of species and such data on the major areas of the brain are also at hand but information on the various subregions is meagre. Such investigations may yet provide clues to the role of lipids in brain function. Compared to CNS, information on PNS is less adequate. Further research on PNS would be worthwhile as it is amenable for experimental manipulation and complex mechanisms such as myelination can be investigated in this tissue. There are reports correlating lipid constituents with the increased complexity in the organization of the nervous system during evolution. This line of investigation may prove useful. The basic aim of research on the lipids of the nervous tissue is to unravel their functional significance.

Metabolism of alpha-terpineol by Pseudomonas incognita

Details of the metabolism of alpha-terpineol by Pseudomonas incognita are presented. Degradation of alpha-terpineol by this organism resulted in the formation of a number of acidic and neutral metabolites. Among the acidic metabolites, beta-isopropyl pimelic acid, 1-hydroxy-4-isopropenyl-cyclohexane-1-carboxylic acid, 8-hydroxycumic acid, oleuropeic acid, cumic acid, and p-isopropenyl benzoic acid have been identified. Neutral metabolites identified were limonene, p-cymene-8-ol, 2-hydroxycineole, and uroterpenol. Cell-free extracts prepared from alpha-terpineol adapted cells were shown to convert alpha-terpineol, p-cymene-8-ol, and limonene to oleuropeic acid, 8-hydroxycumic acid, and perillic acid, respectively, in the presence of NADH. The same cell-free extract contained NAD+ -specific dehydrogenase(s) which converted oleuropyl alcohol, p-cymene-7,8-diol, and perillyl alcohol to their corresponding 7-carboxy acids. On the basis of various metabolites isolated from the culture medium, together with the supporting evidence obtained from enzymatic and growth studies, it appears that P. incognita degrades alpha-terpineol by at least three different routes. While one of the pathways seems to operate via oleuropeic acid, a second may be initiated through the aromatization of alpha-terpineol. The third pathway may involve the formation of limonene from alpha-terpineol and its further metabolism.

Alterations in the activities of the enzymes of proline metabolism in Ragi (Eleusine coracana) leaves during water stress

Free proline content in Ragi (Eleusine coracana) leaves increased markedly (6 to 85 fold) as the degree of water stress, created by polyethylene gylcol treatment, was prolonged There was also a marginal increase in soluble proteins in the stressed leaves as compared to that in the controls. Water stress stimulated the activities of ornithine aminotransferase and pyrroline-5-carboxylate reductase, the enzymes of proline biosynthesis and markedly inhibited the enzymes involved in proline degradation viz., proline oxidase and pyrroline-5-carboxylate dehydrogenase. These results suggest that increase in free proline content of Ragi leaves could be due to enhanced activities of the enzymes synthesizing proline but more importantly due to severe inhibition of the enzymes degrading proline. These observations establish for the first time, the pathway of proline metabolism in plants by way of detection of the activities of all the enzymes involved and also highlight the role of these enzymes in proline accumulation during water stress.