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Query: UMLS:C0026936 (Mycoplasma)
 14,761 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two kinds of arginine deiminase (AD, EC 3.5.3.6) were purified from cell extracts of Mycoplasma arginini (a-AD) and Mycoplasma hominis (h-AD), and their enzymic properties and anti-tumor activities were compared. The a-AD enzyme strongly inhibited the growth of mouse hepatoma cell line MH134 in vitro, and its concentration required for 50% growth inhibition (IC50) was estimated to be about 10 ng/ml. The IC50 value of h-AD against the same cell line was estimated to be about 100 ng/ml, due to its low enzyme activity under the physiological pH condition, i.e., pH 7.4. These results show that the reaction pH profile of the a-AD was superior to that of the h-AD as an anti-tumor enzyme. Moreover, the effects of L-arginine metabolism-related substances on the anti-tumor activity of the a-AD were examined to study the growth-inhibitory mechanism of this enzyme. The addition of 2 or 4 mM L-arginine restored, in a dose-dependent manner, the growth of mouse MH134 hepatoma and Meth A fibrosarcoma cell lines that had been inhibited by 20 ng/ml of the a-AD. The addition of 2 or 4 mM L-ornithine, which is biosynthesized from L-arginine in the urea cycle and is the starting material in the polyamine-biosynthesis pathway, also partially restored it in a dose-dependent manner. These results indicate that the tumor cell growth inhibition caused by a-AD originates from the depletion of the essential nutrient L-arginine, and that the resulting block of the polyamine-biosynthesis pathway is involved in part in the inhibitory mechanism.
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PMID:Anti-tumor activity of arginine deiminase from Mycoplasma argini and its growth-inhibitory mechanism. 759 61

The arginine deiminase (AD) gene was cloned from Mycoplasma arginini and expressed in the cytosol of Escherichia coli as inclusion bodies with an expression level of at least 20% of the total bacterial proteins. The inclusion bodies were solubilized with 6 M guanidine hydrochloride (Gdn-HCl) under reducing conditions, in order to avoid incorrect disulfide-bond formation of the recombinant (r-) AD molecules, and renaturation was performed under various refolding conditions. The optimum renaturation conditions were found to be incubation for 90 h at pH 7.5 and 15 degrees C. The resulting completely refolded r-AD was purified to homogeneity by anion-exchange and arginine-affinity chromatography and its activity yield was 72.5%. The specific activity of the purified r-AD was comparable to and its amino acid composition was identical to those of Mycoplasma AD, and NH2-terminal sequence analysis revealed that its methionine residue corresponding to the translation initiation codon had been removed completely. Anti-tumor activity analyses showed that r-AD inhibited the growth of two mouse cell lines, hepatoma MH134 and fibrosarcoma Meth A, strongly in vitro at concentrations in excess of 10 ng ml-1. Moreover, when MH134-implanted mice were given single intravenous injections of r-AD at doses of 50 mg kg-1 and higher, their survival times were prolonged significantly. These results, taken together, indicate that the enzymatic properties and biological actions of r-AD were highly consistent with those of Mycoplasma AD.
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PMID:High-level expression of Mycoplasma arginine deiminase in Escherichia coli and its efficient renaturation as an anti-tumor enzyme. 776 34

The extent of restriction fragment length polymorphism in genes for the arginine deiminase enzyme among 28 species of mycoplasmas was assessed by Southern blot analysis of DNA digested with EcoRI or TaqI nuclease probed with a 725-bp internal fragment of the arginine deiminase gene from Mycoplasma arginini. The results indicated unexpected heterogeneity among species of a single genus.
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PMID:Polymorphism in genes for the enzyme arginine deiminase among Mycoplasma species. 809 59

Amino acid-degrading enzymes are known to inhibit the growth of tumor cells in culture by depleting amino acids in the medium. Here we demonstrate that arginine deiminase (EC 3.5.3.6) from Mycoplasma arginini had stronger growth-inhibitory activity against all 4 kinds of tumor cell lines tested than L-asparaginase and arginase, which are well-known anti-tumor enzymes. Next, chemical modification of the arginine deiminase molecule with polyethylene glycol was shown to enhance its potency as an anti-tumor enzyme. The percentage of modified amino groups per molecule was estimated to be 51% of the total amino groups, and the average molecular weight was estimated to be about 400,000 by gel-filtration HPLC. The enzymic activity of the modified enzyme was 25.5 units/mg protein, which was equivalent to 57% of that of the native enzyme. The modified enzyme strongly inhibited growth of a mouse hepatoma cell line, MH134, at a concentration of more than 10 ng/ml, showing almost the same dose-response curve as the native enzyme. When a bolus of 5 units of the modified enzyme was intravenously injected into male BDF1 mice, L-arginine in the blood completely disappeared within 5 min, and remained undetectable for more than 8 days. On the other hand, in the case of bolus injection of the same number of units of native enzyme, the plasma L-arginine level recovered up to 66% of the control level at 8 days. These results suggest that this modified enzyme has a longer plasma clearance time and may be more effective as a new anti-tumor agent than the native enzyme.
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PMID:Chemical modification by polyethylene glycol of the anti-tumor enzyme arginine deiminase from Mycoplasma arginini. 827 24

The sequenced genomes of the two closely related bacteria Mycoplasma genitalium and Mycoplasma pneumoniae were compared with emphasis on genome organization and coding capacity. All the 470 proposed open reading frames (ORFs) of the smaller M.genitalium genome (580 kb) were contained in the larger genome (816 kb) of M.pneumoniae. There were some discrepancies in annotation, but inspection of the DNA sequences showed that the corresponding DNA was always present in M. pneumoniae. The two genomes could be subdivided into six segments. The order of orthologous genes was well conserved within individual segments but the order of these segments in both bacteria was different. We explain the different organization of the segments by translocation via homologous recombination. The translocations did not disturb the continuous bidirectional course of transcription in both genomes, starting at the proposed origin of replication. The additional 236 kb in M.pneumoniae,compared with theM.genitalium genome, were coding for 209 proposed ORFs not identified in M.genitalium. Of these ORFs, 110 were specific to M.pneumoniae exhibiting no significant similarity to M.genitalium ORFs, while 76 ORFs were amplifications of ORFs existing mainly as single copies in M. genitalium. In addition, 23 ORFs containing a copy of either one of the three repetitive DNA sequences RepMP2/3, RepMP4 and RepMP5 were annotated in M.pneumoniae but not in M.genitalium,although similar DNA sequences were present. TheM.pneumoniae-specific genes included a restriction-modification system, two transport systems for carbohydrates, the complete set of three genes coding for the arginine dihydrolase pathway and 14 copies of the repetitive DNA sequence RepMP1 which were part of several different translated genes with unknown function.
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PMID:Comparative analysis of the genomes of the bacteria Mycoplasma pneumoniae and Mycoplasma genitalium. 901 18

A growth-inhibitory substance found in the culture of a B-precursor leukemia cell line, NALM-20, was purified from the serum-free culture medium and identified as arginine deiminase derived from Mycoplasma arginini (EC 3.5.3.6). Arginine deiminase strongly inhibited, in a dose-dependent manner, the growth of human T cells and T lymphoblastoid cell lines, but not that of B-precursor and myeloid cell lines. The addition of L-arginine completely restored the growth of T lymphoblastoid cells that had been inhibited by arginine deiminase. The addition of L-ornithine also partially restored it. This enzyme suppressed interleukin-2 (IL-2) production and IL-2 receptor expression in T cells stimulated by non-specific mitogens. The morphologic features of dying cells and DNA fragmentation indicated that arginine deiminase induced apoptotic cell death in T lymphoblasts. Cell cycle analysis revealed that G1-->S transition was blocked in cell treated with arginine deiminase, accompanied by the increase of apoptotic nuclei in the sub-G1 fraction. In conclusion, the deletion of the essential nutrient L-arginine by arginine deiminase significantly inhibited cell growth and activation in T lymphoblasts, accompanied by the induction of apoptotic cell death.
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PMID:Apoptotic cell death of human T lymphoblastoid cells induced by arginine deiminase. 907 17

Mycoplasma infection was detected in cultures of COS cells with a novel, simple assay that detects the conversion of arginine to citrulline by the enzyme, arginine deiminase, specific to all species of mycoplasma. Transfection of COS cells was inhibited in mycoplasma-infected cells, a phenomenon that was readily reversed by treatment with a mycoplasma removal agent. Cultures of cells used for transfection should be regularly monitored for evidence of mycoplasma by assay of arginine deiminase activity or by other methods.
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PMID:Detection of mycoplasma infection of mammalian cells. 914 78

Mollicutes or mycoplasmas are a class of wall-less bacteria descended from low G + C% Gram-positive bacteria. Some are exceedingly small, about 0.2 micron in diameter, and are examples of the smallest free-living cells known. Their genomes are equally small; the smallest in Mycoplasma genitalium is sequenced and is 0.58 mb with 475 ORFs, compared with 4.639 mb and 4288 ORFs for Escherichia coli. Because of their size and apparently limited metabolic potential, Mollicutes are models for describing the minimal metabolism necessary to sustain independent life. Mollicutes have no cytochromes or the TCA cycle except for malate dehydrogenase activity. Some uniquely require cholesterol for growth, some require urea and some are anaerobic. They fix CO2 in anaplerotic or replenishing reactions. Some require pyrophosphate not ATP as an energy source for reactions, including the rate-limiting step of glycolysis: 6-phosphofructokinase. They scavenge for nucleic acid precursors and apparently do not synthesize pyrimidines or purines de novo. Some genera uniquely lack dUTPase activity and some species also lack uracil-DNA glycosylase. The absence of the latter two reactions that limit the incorporation of uracil or remove it from DNA may be related to the marked mutability of the Mollicutes and their tachytelic or rapid evolution. Approximately 150 cytoplasmic activities have been identified in these organisms, 225 to 250 are presumed to be present. About 100 of the core reactions are graphically linked in a metabolic map, including glycolysis, pentose phosphate pathway, arginine dihydrolase pathway, transamination, and purine, pyrimidine, and lipid metabolism. Reaction sequences or loci of particular importance are also described: phosphofructokinases, NADH oxidase, thioredoxin complex, deoxyribose-5-phosphate aldolase, and lactate, malate, and glutamate dehydrogenases. Enzymatic activities of the Mollicutes are grouped according to metabolic similarities that are taxonomically discriminating. The arrangements attempt to follow phylogenetic relationships. The relationships of putative gene assignments and enzymatic function in My. genitalium, My. pneumoniae, and My. capricolum subsp. capricolum are specially analyzed. The data are arranged in four tables. One associates gene annotations with congruent reports of the enzymatic activity in these same Mollicutes, and hence confirms the annotations. Another associates putative annotations with reports of the enzyme activity but from different Mollicutes. A third identifies the discrepancies represented by those enzymatic activities found in Mollicutes with sequenced genomes but without any similarly annotated ORF. This suggests that the gene sequence is significantly different from those already deposited in the databanks and putatively annotated with the same function. Another comparison lists those enzymatic activities that are both undetected in Mollicutes and not associated with any ORF. Evidence is presented supporting the theory that there are relatively small gene sequences that code for functional centers of multiple enzymatic activity. This property is seemingly advantageous for an organism with a small genome and perhaps under some coding restraint. The data suggest that a concept of "remnant" or "useless genes" or "useless enzymes" should be considered when examining the relationship of gene annotation and enzymatic function. It also suggests that genes in addition to representing what cells are doing or what they may do, may also identify what they once might have done and may never do again.
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PMID:The comparative metabolism of the mollicutes (Mycoplasmas): the utility for taxonomic classification and the relationship of putative gene annotation and phylogeny to enzymatic function in the smallest free-living cells. 943 86

Streptococcus pyogenes (group A Streptococcus) cell extracts (CE) have a remarkably powerful and dose-dependent inhibitory effect on antigen, superantigen, or mitogen-stimulated human peripheral blood mononuclear cell (PBMC) proliferation in vitro. Purification of the inhibitory component present in S. pyogenes type M5 (Manfredo strain) CE by anion-exchange chromatography followed by gel filtration chromatography showed that the inhibitor had an approximate native molecular mass of 100 kDa. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified inhibitory fractions followed by silver staining gave a single band with an approximate molecular mass of 47 kDa, indicating that the inhibitor is composed of two identical subunits. NH2-terminal sequencing of the protein revealed that it was identical to the previously characterized streptococcal acid glycoprotein (SAGP); this protein possesses between 31.5 and 39.0% amino acid identity with arginine deiminase (AD) from Mycoplasma hominis, Mycoplasma arginini, Pseudomonas putida, and Pseudomonas aeruginosa. AD enzyme activity was present in unfractionated CE prepared from a range of streptococcal strains, and partially purified inhibitory fractions of Manfredo CE also had high levels of activity. The inhibitory effect of Manfredo CE was overcome by the addition of L-arginine to proliferation assays in which human PBMC were stimulated with phytohemagglutinin. We conclude that SAGP, or its homolog, possesses AD activity and that the potent inhibition of proliferation of human T cells by streptococcal CE is due to activity of this enzyme.
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PMID:Inhibition of human peripheral blood mononuclear cell proliferation by Streptococcus pyogenes cell extract is associated with arginine deiminase activity. 963 65

We used confluent cultures of dog gallbladder epithelial cells, stimulated by conditioned medium from a culture of human neonatal foreskin fibroblasts, to establish the presence of inducible nitric oxide synthase (NOS, EC 1.14.13.39). Assay was by conversion of radiolabeled arginine to citrulline. By 4 days after addition of the conditioned medium, a relatively high level of activity was observed. However, further study showed that the enzyme did not require addition of the usual cofactors for maximal activity (NADPH, FAD, FMN and tetrahydrobiopterin) and was stable in the absence of anti-proteolytic agents. Our suspicion that this enzyme might not be NOS but arginine deiminase (EC 3.5.3.6) was confirmed by enzyme purification and by the liberation of ammonia during enzyme reaction. This enzyme, which is absent from primates and virtually confined to single-cell organisms, suggested the presence of Mycoplasma, a common contaminant of cell cultures, and it was subsequently confirmed that the fibroblast culture was a source of Mycoplasma. With the widespread interest in nitric oxide and NOS, and common use of the convenient [3H]arginine assay, there is a considerable danger of the two enzymes being confused. At the very least, it is necessary to check for activity in the absence of added cofactors.
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PMID:Caveat: mycoplasma arginine deiminase masquerading as nitric oxide synthase in cell cultures. 973 59


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