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At least 19 recordsLinked to original sources
PubMed2004

Ion chromatographic quantification of cyanate in urea solutions: estimation of the efficiency of cyanate scavengers for use in recombinant protein manufacturing.

The chaotrope urea is commonly used during recombinant protein manufacturing as a denaturant/solublizing agent. The adventitious accumulation of cyanate in urea solutions during product manufacturing can cause unwanted carbamylation of proteins, leading to alterations in drug product structure, stability and function. We have developed an ion chromatographic method to quantify cyanate production in urea solutions, suitable for analysis of samples from manufacturing process buffers. We discuss assay development, system suitability criteria and limitations on assay applicability. The assay has a linear range from 2 to 250 microM, with LOQ/LOD values of 6 and 2 microM, respectively. Assay accuracy through spike/recovery testing were established and both precision and intermediate precision were estimated. We assessed the utility of the assay by testing a variety of biological buffers and potential cyanate scavengers, which could be used during protein purification processes, for their ability to control the level of cyanate in 8 M urea solutions buffered over the range of pH 5-10. Our results demonstrate pH dependence for prevention of cyanate accumulation by these buffers/scavengers and indicate useful buffers, pH ranges, and additives for controlling cyanate accumulation during recombinant protein manufacturing. The pertinence of these approaches in preventing protein carbamylation during manufacturing are discussed.

Chromatography, Liquid↗
PubMed1975

Bioavailability of sodium cyanate in patients with sickle cell disease and the lack of inhibition in vitro of globin synthesis at in vivo concentrations of cyanate.

Studies have been made on the bioavailability in blood of sodium cyantate administered orally in gelatin capsules, in gelatin capsules plus antacid, in enteric-coated capsules and in cocoa butter suppositories administered rectally to patients with sickle cell disease. Maximal blood concentrations of cyanate did not exceed 0.4 mM. Sodium cyanate taken orally in gelatin capsules yielded the highest blood concentrations of the drug, but the peak concentrations and curve areas were not necessarily dose-related. The duration of the drug in the circulation was about 210 minutes. Administration of sodium cyanate in the gelatin capsules, taken with an antacid, improved the dose-response relationship within a given patient. Enteric-coated capsules and suppositories were found to show variable and low bioavailability profiles, respectively. Variability in bioavailability between patients with a given dosage form requires further study. Since the concentration of cyanate attainable in vivo does not inhibit synthesis of either the alpha- or beta-chain of hemoglobin in vitro, previous reports on the inhibitory effects of 10 to 100 mM cyanate on globin synthesis in vitro do not appear to be relevant.

Anemia, Sickle Cell↗
PubMed1986

Bendazac prevents cyanate binding to soluble lens proteins and cyanate-induced phase-separation opacities in vitro: a possible mechanism by which bendazac could delay cataract.

The reaction of lens proteins with cyanate (carbamylation) causes many changes seen in human cataract including disruption of the protein conformations. Bendazac, a putative anti-cataract drug, decreases the binding of cyanate to lens proteins and prevents the cyanate-induced elevation of the phase separation temperature in incubated rat lenses. Its major metabolite, 5-hydroxybendazac, also inhibits the binding of cyanate to lens proteins even when it is present only during a pre-incubation period. The metabolite is more effective than the parent compound.

Animals↗
PubMed1975

In vivo hepatic and intestinal toxicity of sodium cyanate in rats: cyanate-induced alterations in hepatic glycogen metabolism.

To determine the hepatic and intestinal toxicity of sodium cyanate, this compound was administered to rats by orogastric tube (PO) or intraperitoneal injection (IP). At low dosage (50 mg. per kilogram per day PO for 8 weeks), the animals showed no clinical effects other than mild lethargy. They had normal intestinal absorption studies, but demonstrated decreased liver G6PD activity and a slight increase in hepatic glycogen. At higher dose levels (200 mg. per kilogram per day PO for 10 days, 400 mg. per kilogram per day PO for 3 days, and 100 mg. per kilogram per day IP for 10 days), the animals became very lethargic and developed hind-limb paralysis; many animals died during the period of dosing. The severity and rate of onset of symptoms increased proportionally with the dose level. Liver sections from rats receiving these higher doses showed striking increases in glycogen deposition. Activities of hepatic enzymes involved in glycogen synthesis and degradation were measured in rats receiving 200 mg. per kilogram per day PO or 100 mg. per kilogram per day IP. Significant decreases were noted in the activities of glucose-6-phosphatase and G6PD in PO-dosed rats. The activities of phosphorylase, UDPG-pyrophosphorylase, glycogen synthetase, phosphoglucomutase, and debrancher did not differ from control rats. In IP-dosed rats, significant decreases were observed in the activities of glucose-6-phosphatase, G6PD, phosphorylase, and UDPG-pyrophosphorylase, but not in the other glycogen-related enzymes. Our data suggest that sodium cyanate affects several enzymes of hepatic glycogen metabolism but that the enzymes vary in their susceptibility (glucose-6-phosphatase and G6PD greater than phosphorylase and UDPG pyrophosphorylase.

Animals↗
PubMed2004

Potassium hexacyanoferrate(II)--a new cyanating agent for the palladium-catalyzed cyanation of aryl halides.

A new advantageous cyanating agent, potassium hexacyanoferrate(II), is described for the palladium-catalyzed cyanation of aryl halides. All cyanide ions on the iron(II) center can be transferred to the aryl halide using palladium(II) acetate and dppf as the catalyst. Under optimized reaction conditions good yields of benzonitriles and unprecedented catalyst productivities are observed.

Journal Article↗
PubMed1993

A physiological role for cyanate-induced carbonic anhydrase in Escherichia coli.

Cyanate induces expression of the cyn operon in Escherichia coli. The cyn operon includes the gene cynS, encoding cyanase, which catalyzes the reaction of cyanate with bicarbonate to give ammonia and carbon dioxide. A carbonic anhydrase activity was recently found to be encoded by the cynT gene, the first gene of the cyn operon; it was proposed that carbonic anhydrase prevents depletion of bicarbonate during cyanate decomposition due to loss of CO2 by diffusion out of the cell (M. B. Guilloton, J. J. Korte, A. F. Lamblin, J. A. Fuchs, and P. M. Anderson, J. Biol. Chem. 267:3731-3734, 1992). The function of the product of the third gene of this operon, cynX, is unknown. In the study reported here, the physiological roles of cynT and cynX were investigated by construction of chromosomal mutants in which each of the three genes was rendered inactive. The delta cynT chromosomal mutant expressed an active cyanase but no active carbonic anhydrase. In contrast to the wild-type strain, the growth of the delta cynT strain was inhibited by cyanate, and the mutant strain was unable to degrade cyanate and therefore could not use cyanate as the sole nitrogen source when grown at a partial CO2 pressures (pCO2) of 0.03% (air). At a high pCO2 (3%), however, the delta cynT strain behaved like the wild-type strain; it was significantly less sensitive to the toxic effects of cyanate and could degrade cyanate and use cyanate as the sole nitrogen source for growth. These results are consistent with the proposed function for carbonic anhydrase. The chromosomal mutant carrying cynS::kan expressed induced carbonic anhydrase activity but no active cyanase. The cynS::kan mutant was found to be much less sensitive to cyanate than the delta cynT mutant at a low pCO2, indicating that bicarbonate depletion due to the reaction of bicarbonate with cyanate catalyzed by cyanase is more deleterious to growth than direct inhibition by cyanate. Mutants carrying a nonfunctional cynX gene (cynX::kan and delta cynT cynX::kan) did not differ from the parental strains with respect to cyanate sensitivity, presence of carbonic anhydrase and cyanase, or degradation of cyanate by whole cells; the physiological role of the cynX product remains unknown.

Bicarbonates↗
PubMedDate not supplied

Chronic peritoneal inflammation by cyanate in rats.

OBJECTIVE: During peritoneal dialysis, the peritoneum is exposed to waste products, including urea. Urea forms cyanate spontaneously at body temperature and pH, and cyanate carbamylates amino acids, peptides, and proteins. Cyanate may contribute to peritoneal injury with morphological changes in the peritoneum. To test this hypothesis, we injected cyanate into rats. METHODS: Experiments were performed in two groups of 7 rats each. In the cyanate group, each rat received 1 mL of 1.5 micromol/L potassium cyanate dissolved in 40 mmol/L sodium bicarbonate solution intraperitoneally each experiment day. In the control group, each rat received 1 mL of 1.5 micromol/L potassium bicarbonate instead of potassium cyanate. The rats in both groups were anesthetized and killed at the 85th day after the first injection. After formalin fixation, tissue samples from abdominal walls and livers were sliced, embedded in a standard manner, and stained with hematoxylin and eosin. RESULTS: Parietal peritoneum from rats in the cyanate group showed a mild increase in the number of fibroblasts, with collagen deposits, infiltration by mononuclear cells, vascular congestion, round-shaped transformation of mesothelial cells, widening of submesothelial spaces, and abundant denudation of mesothelial cells. The visceral peritoneum from rats in the cyanate group showed collagen deposits with fibroblastic proliferation. CONCLUSIONS: Cyanate can induce chronic inflammation in the peritoneum, and exposure of the peritoneum to cyanate may contribute to peritoneal injury in patients being treated with peritoneal dialysis.

Animals↗
PubMed2000

Impaired biological activity of erythropoietin by cyanate carbamylation.

AIMS: During advanced renal failure, particularly in patients with end-stage renal disease (ESRD), proteins are carbamylated as a result of a reaction with cyanate. Some or all of the cyanate is derived from urea. If the carbamylation of proteins adversely alters their biologic activities, then urea must be viewed as an uremic toxin, rather than a surrogate. Therefore, we studied the effect of cyanate carbamylation on the erythropoietic activity of erythropoietin (EPO) in a rodent model. METHODS: EPO was carbamylated by incubation with cyanate at 37 degrees C. The extent of carbamylation was monitored using trinitrobenzenesulfonic acid. In Sprague-Dawley rats the erythrocyte count, hemoglobin concentration, and hematocrit were measured after the twice-weekly subcutaneous injection of either EPO or carbamylated EPO for 3 weeks. Two additional control groups received physiologic saline or 0.2 ml of 1 M cyanate. RESULTS: The level of carbamylated EPO was increased as the time of exposure to cyanate increased from 1 to 6 h, and as the cyanate concentration increased from 8 to 2,000 mM. EPO injections caused significantly large increases in all erythropoietic measures. Physiologic saline or 1 M cyanate-injected controls and the carbamylated EPO-injected animals demonstrated no change from baseline in erythropoietic parameters. CONCLUSION: These results support that EPO exposed to high levels of cyanate in vitro demonstrates diminished biologic activity in healthy Sprague-Dawley rats. This effect may be manifested by the carbamylation of EPO by the cyanate. Should this occur in ESRD patients, it may contribute to the suboptimal erythropoietic response to EPO therapy associated with high urea levels, especially related to inadequate dialysis. Targeting dialysis doses specifically to urea concentrations may be more important than previously considered.

Animals↗
PubMed1975

Cyanate modification of essential lysyl residues of the diphosphopyridine nucleotide-specific isocitrate dehydrogenase of pig heart.

The DPN-specific isocitrate dehydrogenase of pig heart is totally and irreversibly inactivated by 0.05 M potassium cyanate at pH 7.4 A plot of the rate constant versus cyanate concentration is not linear, but rather exhibits saturation kinetics, implying that cyanate may bind to the enzyme to give an enzyme-cyanate complex (K equal 0.125 M) prior to the covalent reaction. In the presence of manganous ion the addition of isocitrate protects the enzyme against cyanate inactivation, indicating that chemical modification occurs in the active site region of the enzyme. The dependence of the decrease of the rate constant for inactivation on the isocitrate concentration yields a dissociation constant for the enzyme-manganese-isocitrate complex which agrees with the Michaelis constant. The allosteric activator ADP, which lowers the Michaelis constant for isocitrate, does not itself significantly affect the cyanate reaction; however, it strikingly enhances the protection by isocitrate. The addition of the chelator EDTA essentially prevents protection by isocitrate and manganous ion, demonstrating the importance of the metal ion in this process. The substrate alpha-ketoglutarate and the coenzymes DPN and DPNH do not significantly affect the rate of modification of the enzymes by cyanate. Incubation of isocitrate dehydrogenase with 14C-labeled potassium cyanate leads to the incorporation of approximately 1 mol of radioactive cyanate per peptide chain concomitant with inactivation. Analysis of acid hydrolysates of the radioactive enzyme reveals that lysyl residues are the sole amino acids modified. These results suggest that cyanate, or isocyanic acid, may bind to the active site of this enzyme as an analogue of carbon dioxide and carbamylate a lysyl residue at the active site.

Adenosine Diphosphate↗
PubMed1993

Alteration of the systemic antitumor activity of melphalan by sodium cyanate in MOPC-460D myeloma-bearing BALB/c mice.

Sodium cyanate is a selective inhibitor of protein synthesis in a variety of mammalian tumor cells without a corresponding effect on normal tissue of the tumor-bearing animals. In the present study, we investigated the potential role of sodium cyanate in the augmentation of the antitumor activity of melphalan in MOPC-460D myeloma-bearing BALB/c mice. The simultaneous intraperitoneal injection of sodium cyanate, 250 mg/kg, and melphalan, 12 mg/kg, followed by another dose of sodium cyanate, 200 mg/kg, administered 18 hours later, resulted in a tumor growth inhibition index (TGII) of 207%. In contrast, melphalan or sodium cyanate administered separately at the same dose induced a TGII of 133% and 15%, respectively, when compared to control animals. Furthermore, a direct comparison of the volume of tumor implants in mice treated with the combination of sodium cyanate and melphalan vs. those treated with melphalan alone showed a statistically significant growth inhibition in favor of the sodium cyanate and melphalan combination on days 35, 39, and 42 from initiation of treatment. The data presented here suggest that the antitumor activity of melphalan could be increased, with moderate toxicity, by the concomitant intraperitoneal administration of sodium cyanate in BALB/c mice bearing measurable subcutaneous MOPC-460D tumor transplants. This is the first report of an increase in melphalan antitumor activity by sodium cyanate at a tumor location distant from the site of injection.

Animals↗
PubMed1998

A novel pink-pigmented facultative methylotroph, Methylobacterium thiocyanatum sp. nov., capable of growth on thiocyanate or cyanate as sole nitrogen sources.

The isolation and properties of a novel species of pink-pigmented methylotroph, Methylobacterium thiocyanatum, are described. This organism satisfied all the morphological, biochemical, and growth-substrate criteria to be placed in the genus Methylobacterium. Sequencing of the gene encoding its 16S rRNA confirmed its position in this genus, with its closest phylogenetic relatives being M. rhodesianum, M. zatmanii and M. extorquens, from which it differed in its ability to grow on several diagnostic substrates. Methanol-grown organisms contained high activities of hydroxypyruvate reductase -3 micromol NADH oxidized min-1 (mg crude extract protein)-1], showing that the serine pathway was used for methylotrophic growth. M. thiocyanatum was able to use thiocyanate or cyanate as the sole source of nitrogen for growth, and thiocyanate as the sole source of sulfur in the absence of other sulfur compounds. It tolerated high concentrations (at least 50 mM) of thiocyanate or cyanate when these were supplied as nitrogen sources. Growing cultures degraded thiocyanate to produce thiosulfate as a major sulfur end product, apparently with the intermediate formation of volatile sulfur compounds (probably hydrogen sulfide and carbonyl sulfide). Enzymatic hydrolysis of thiocyanate by cell-free extracts was not demonstrated. Cyanate was metabolized by means of a cyanase enzyme that was expressed at approximately sevenfold greater activity during growth on thiocyanate [Vmax 634 +/- 24 nmol NH3 formed min-1 (mg protein)-1] than on cyanate [89 +/- 9 nmol NH3 min-1 (mg protein)-1]. Kinetic study of the cyanase in cell-free extracts showed the enzyme (1) to exhibit high affinity for cyanate (Km 0.07 mM), (2) to require bicarbonate for activity, (3) to be subject to substrate inhibition by cyanate and competitive inhibition by thiocyanate (Ki 0.65 mM), (4) to be unaffected by 1 mM ammonium chloride, (5) to be strongly inhibited by selenocyanate, and (6) to be slightly inhibited by 5 mM thiosulfate, but unaffected by 0.25 mM sulfide or 1 mM thiosulfate. Polypeptides that might be a cyanase subunit (mol.wt. 17.9 kDa), a cyanate (and/or thiocyanate) permease (mol.wt. 25.1 and 27.2 kDa), and a putative thiocyanate hydrolase (mol.wt. 39.3 kDa) were identified by SDS-PAGE. Correlation of the growth rate of cultures with thiocyanate concentration (both stimulatory and inhibitory) and the kinetics of cyanase activity might indicate that growth on thiocyanate involved the intermediate formation of cyanate, hence requiring cyanase activity. The very high activity of cyanase observed during growth on thiocyanate could be in compensation for the inhibitory effect of thiocyanate on cyanase. Alternatively, thiocyanate may be a nonsubstrate inducer of cyanase, while thiocyanate degradation itself proceeds by a carbonyl sulfide pathway not involving cyanate. A formal description of the new species (DSM 11490) is given.

Cyanates↗
PubMed2004

Inhibition of erythropoietin activity by cyanate.

OBJECTIVE: Increased urea concentration is a measure of advanced renal failure and the adequacy of renal replacement therapy in end-stage renal disease (ESRD). Altered biologic activity due to changes in protein structure occurs when cyanate, formed spontaneously from urea, reacts with proteins. Carbamylation results in impaired erythropoietin (EPO) activity when high concentrations of cyanate react with EPO. In this study, the activity of carbamylated EPO (C-EPO), formed at a cyanate concentration which may occur in vivo, was studied in Sprague-Dawley rats. MATERIAL AND METHODS: The extent of carbamylation, causing loss of free amino groups, was monitored using trinitrobenzenesulfonic acid. Erythrocyte, hemoglobin, hematocrit and leukocyte levels were measured after either EPO, incubated EPO, C-EPO, physiologic saline or cyanate (1.5 microM; 0.2 ml) were injected subcutaneous twice weekly for 3 weeks in rats. RESULTS: In vitro carbamylation of EPO was time- and concentration-dependent. C-EPO concentration increased as the duration of exposure to cyanate increased from 6 to 72 h, or as cyanate concentration increased from 15 nM to 1.5 microM. Injections of EPO caused significant increases in vivo in all erythropoietic measures. In contrast, injections of C-EPO, physiologic saline or 1.5 microM cyanate caused no change from baseline. CONCLUSIONS: These results demonstrated diminished biologic activity in healthy rats by C-EPO formed in vitro at cyanate concentrations that may be found in vivo. C-EPO and high urea-derived cyanate levels may contribute to suboptimal erythropoietic responses to EPO therapy for chronic renal failure and ESRD, and may provide another measurement indicating inadequate dialysis.

Animals↗
PubMed1995

Role of bicarbonate/CO2 in the inhibition of Escherichia coli growth by cyanate.

Cyanase is an inducible enzyme in Escherichia coli that catalyzes the reaction of cyanate with bicarbonate to give two CO2 molecules. The gene for cyanase is part of the cyn operon, which includes cynT and cynS, encoding carbonic anhydrase and cyanase, respectively. Carbonic anhydrase functions to prevent depletion of cellular bicarbonate during cyanate decomposition (the product CO2 can diffuse out of the cell faster than noncatalyzed hydration back to bicarbonate). Addition of cyanate to the culture medium of a delta cynT mutant strain of E. coli (having a nonfunctional carbonic anhydrase) results in depletion of cellular bicarbonate, which leads to inhibition of growth and an inability to catalyze cyanate degradation. These effects can be overcome by aeration with a higher partial CO2 pressure (M. B. Guilloton, A. F. Lamblin, E. I. Kozliak, M. Gerami-Nejad, C. Tu, D. Silverman, P. M. Anderson, and J. A. Fuchs, J. Bacteriol. 175:1443-1451, 1993). The question considered here is why depletion of bicarbonate/CO2 due to the action of cyanase on cyanate in a delta cynT strain has such an inhibitory effect. Growth of wild-type E. coli in minimal medium under conditions of limited CO2 was severely inhibited, and this inhibition could be overcome by adding certain Krebs cycle intermediates, indicating that one consequence of limiting CO2 is inhibition of carboxylation reactions. However, supplementation of the growth medium with metabolites whose syntheses are known to depend on a carboxylation reaction was not effective in overcoming inhibition related to the bicarbonate deficiency induced in the delta cynT strain by addition of cyanate. Similar results were obtained with a deltacyn strain (since cyanase is absent, this strain does not develop a bicarbonate deficiency when cyanate is added); however, as with the deltacynT strain, a higher partial CO(2) pressure in the aerating gas or expression of carbonic anhydrase activity (which contributes to a higher intercellular concentration of bicarbonate/CO(2)) significantly reduced inhibition of growth. There appears to be competition between cyanate and bicarbonate/CO(2) at some unknown but very important site such that cyanate binding inhibits growth. These results suggest that bicarbonate/CO(2) plays a significant role in the growth of E. coli other than simply as a substrate for carboxylation reactions and that strains with mutations in the cyn operon provide a unique model system for studying aspects of the metabolism of bicarbonate/CO(2) and its regulation in bacteria.

Bicarbonates↗
PubMed1975

Cyanate as an inactivator of complement proteins.

Sodium cyanate added to normal human serum or serum from patients with sickle-cell disease resulted in the functional inactivation of C3, C5, C6, C7, and the C3b inactivator, but not C8 and C9. Final concentrations as low as 0.5 mM in serum caused inactivation of 12 to 64% of the C3 after 8 hr at 37 degrees C. The activity of the inactivated C3, C5, and C3b inactivator was not restored by dialysis. Most of the functional activity of C3 in cyanate-treated sera was destroyed by very small quantities of 14C-labeled cyanate that was bound to the protein. C3 inactivation by cyanate occurred in heated sera (50 degrees C, 30 min) and sera treated with EDTA, probably indicating that one mechanism for inactivation was by a direct carbamylation reaction. Both C3 and C5 showed two anodal-migrating forms in two dimensional antigen-antibody crossed electrophoresis in some sera treated with low concentrations of cyanate. Measurements of circular dichroism of highly purified carbamylated C3 showed no detectable changes in structure even though most of the functional activity was destroyed. Purified, inactive C3 that was carbamylated with 14C-labeled cyanate was capable of binding to EAC142, but the resulting EAC1423 was weakly positive for immune adherence and negative for agglutination with anti-C3 antiserum. Unlabeled, cell-bound C3b on EAC142 was not susceptible to cyanate action as shown by no loss in immune adherence and positive agglutination with anti-C3 antiserum. The C3b inactivator was more susceptible to cyanate than C3 in a short time period, whereas both were inactivated after 8 hr. Since cyanate is currently being evaluated as a treatment for sickle-cell disease, the inactivation of C3 by the drug is an important consideration for such patients who are already deficient in C3 dependent heat-labile opsonins that aid in host defense.

Anemia, Sickle Cell↗
PubMed1999

Sodium cyanate alters glutathione homeostasis in rodent brain: relationship to neurodegenerative diseases in protein-deficient malnourished populations in Africa.

Sodium cyanate, a neurotoxic chemical in rodents, primates and humans, is implicated in neurodegenerative disorders in protein-deficient populations subsisting in parts of Africa on the cyanogenic plant cassava. The molecular and cellular mechanisms of cyanate neurotoxicity are not understood. This study investigates the effect of sodium cyanate on glutathione (GSH) homeostasis in rodent brain and liver in vitro and in vivo. GSH levels in mouse brain were rapidly, time- and dose-dependently decreased following intraperitoneal administration of 100, 200 or 300 mg/kg sodium cyanate. By contrast, GSH disulfide (GSSG) levels were increased and GSH/GSSG ratios were decreased in a dose-dependent manner in rat brain. Sodium cyanate depleted GSH levels in all regions of mouse brain. Brain glutathione reductase activity was dose-dependently inhibited, while glutathione peroxidase activity was not affected by sodium cyanate. The disruption of GSH homeotasis, as evidenced by reduced tissue GSH/GSSG ratios, likely results from cyanate-induced inhibition of glutathione reductase activity. The results of this study suggest that cyanate neurotoxicity, and perhaps cassava-associated neurodegenerative diseases, are mediated in part by disruption of glutathione homeostasis in neural tissue.

Africa↗
PubMed1990

The cyanase operon and cyanate metabolism.

Cyanase is an inducible enzyme in E. coli that catalyzes bicarbonate-dependent decomposition of cyanate. It is encoded as part of an operon we have named the cyn operon, which includes three genes in the following order: cynT (cyanate permease), cynS (cyanase), and cynX (protein of unknown function). The direction of transcription is opposite to that of the lac operon, and the 3'-end of the cyn operon overlaps the 3'-end of the lac operon by 98 nucleotides. The gene cynR (regulatory protein) is located upstream from the cyn operon, and its transcription is opposite that of the cyn operon. The genes of the cyn operon and the cynR gene have been cloned, sequenced and over-expressed. Cyanate at concentrations of about 1 mM is toxic to strains of E. coli lacking the cyanase gene, but strains in which the inducible gene for cyanase is present can grow on cyanate as the sole source of nitrogen at concentrations as high as 20 mM. The presence of cyanase itself is not sufficient to overcome cyanate toxicity--the permease must also be present. Strains lacking the cyanase gene, but having a functional permease gene, are extremely sensitive to cyanate. Uptake of cyanate involves the product of the permease gene in an energy-dependent process. It appears that the cyn operon has evolved to function in detoxification/decomposition of cyanate arising from both intra- and extracellular sources.

Aminohydrolases↗