How does ciprofloxacin inhibit dna gyrase

DNA gyrase has been known for some time to exist in plants 6 , 7 , but the lethality of knockouts and difficulties in producing active recombinant protein have meant that plant gyrase has remained largely uncharacterized. However, this had not been established.

Further it has been shown that A. The most direct way to ascertain the target of a drug is to isolate mutants that map to the gene encoding the target enzyme.

We used EMS-mutagenized A. This mutation has not been reported in E. However, it is known that the same mutation leads to ciprofloxacin resistance in other species, e. Treatment of A. With wild-type plants, many of the chloroplasts appeared to be in the process of dividing, implying that the drug was interrupting organellar replication, as might be predicted. Interestingly, we found little difference in mitochondrial morphology for wild-type plants in the presence of ciprofloxacin, suggesting that gyrase may not play such a crucial role in these organelles.

These results further endorse the view that gyrase is targeted to chloroplasts and is targeted by ciprofloxacin 6 , 7. We applied two further tests to prove that DNA gyrase is active in A. Moreover when we expressed the AtGyrA AV mutant protein, we found that it conferred drug resistance.

Taken together these experiments confirm that DNA gyrase is active in A. The difficulties in heterologously expressing the A. It is hoped that the results described in this study, particularly the successful heterologous expression in insect cells, will pave the way for future work on this enzyme. In particular, it would be useful to ascertain whether AtGyrB1 and AtGyrB2 have distinct roles and whether both are able to complex with AtGyrA to constitute active supercoiling enzymes.

The confirmation that A. All authors reviewed the results and approved and edited the final version of the manuscript. This work was also was supported by a Ph.

The authors declare that they have no conflicts of interest with the contents of this article. National Center for Biotechnology Information , U. Journal List J Biol Chem v. J Biol Chem. Published online Dec 9. Katherine M. Lesley A. Melisa K. Joshua S.

Author information Article notes Copyright and License information Disclaimer. Received Sep 9; Revised Dec 8. This article has been cited by other articles in PMC. Open in a separate window. Mutagenesis EMS-mutagenized A. No drug Gluf. Reconstitution of A. Discussion DNA gyrase has been known for some time to exist in plants 6 , 7 , but the lethality of knockouts and difficulties in producing active recombinant protein have meant that plant gyrase has remained largely uncharacterized.

Author Contributions A. References 1. Bush N. EcoSal Plus 6 , Vos S. Cell Biol. Gellert M. Collin F. Pommier Y. Cho H. Wall M. Dar M. Evans-Roberts K. Burlison J. Robinson M. Aoyama T. Plant J.

Waterhouse P. Jones J. Bechtold N. III , — [ Google Scholar ]. Neff M. Trends Genet. Kreuzer K. Hallett P. Gene 93 , — [ PubMed ] [ Google Scholar ]. S and R represent supercoiled and relaxed pUC18, respectively. Cleavage reactions were carried out with M. Cleavage reactions performed in the presence of different concentrations of CcdB with a E.

Lane M contains a 1 kb ladder Life Technologies. Relative cleavage is the amount of cleavage product seen in the presence of CcdB normalized to the intrinsic cleavage produced by the enzyme alone. Corresponding author. The authors thank J. Wang and A. Maxwell for overexpressing constructs of E. We also acknowledge R. Goetschi for cyclothialidines, R. Varadarajan for CcdB and K. Muniyappa for etoposide. Radha is acknowledged for technical assistance.

This work is supported by research grants from the Departments of Science and Technology, and Biotechnology, Government of India. Gellert, M. Reece, R. DNA gyrase: structure and function. Wang, J. Quarterly Review of Biophysics 31 , — Hooper, D. Bacterial topoisomerases, anti-topoisomerases, and anti-topoisomerase resistance.

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As an inhibitor of these enzymes, ciprofloxacin is able to effectively disrupt replication, ultimately leading to apoptosis of the replicating cell. Based on the patient's illness, this antibiotic was prescribed at a therapeutic dosage that would achieve maximal inhibition of these topoisomerase enzymes.

Active ciprofloxacin molecules bind two sites within the topoisomerase and are held in place through Van der Waals forces between the drug's ketone group and a magnesium ion. The presence of this magnesium ion allows for the formation of an octahedral coordination sphere at the carbonyl and carboxylic acid groups attached to C3 and C4 respectively, as well as hydrogen bonding to two amino acid residues that attach the drug to the topoisomerase. Once bound, the drug acts like a physical wedge within topoisomerase and prevents it from binding DNA and exerting its effects during replication.

However, the use of this antibiotic may cause unintentional drug-drug interactions DDIs when used in combination with other medications due to its metabolism by the Cytochrome P CYP family of enzymes. As previously mentioned, ciprofloxacin is an inhibitor of the CYP1A2 enzyme; the same enzyme necessary for the metabolism of the muscle-relaxant, tizanidine.

As a centrally acting alpha-2 adrenergic agonist, tizanidine suppresses activity in the sympathetic nervous system and may lead to effects such as decreased heart rate and reduced blood pressure in addition to other central nervous system effects such as drowsiness, fatigue, and GI upset. Due to the inhibition of CYP1A2, tizanidine is metabolized more slowly and with subsequent dosing, may accumulate in the blood.

It is this accumulation that increases the risk of occurrence for serious side effects such as hypotension and sedation as seen with the patient. Due to the appearance of these serious side effects, it became necessary to change this patient's therapy in order to treat the bacterial infection while avoiding any potential harm as the result of the DDI.

In addition, abrupt discontinuation of tizanidine may result in negative cardiovascular effects such as rebound hypertension, tachycardia, and hypertonia. For this reason it is suggested that tizanidine be tapered down as opposed to discontinued abruptly. After a brief discussion with the physician, the patient was administered isotonic intravenous fluids to restore a normotensive state and was able to speak to the pharmacist about the cause of the new symptoms they had been experiencing.

This medication was given to replace their previous tizanidine regimen. The patient was strongly advised against alcohol use in combination with diazepam and was educated on the potential side effects including trouble sleeping, headache, and dizziness.

By instituting this 'taper', the patient would be able to fully discontinue the tizanidine after 3 days and avoid the negative cardiovascular effects that are common when abruptly stopping a2-agonists. This new treatment regimen was developed through inter-professional cooperation utilizing pharmacodynamic profiles 9, 10, 11, 12, 13 and drug interaction reports based on the patient's current need for infrequent muscle spasm relief as well as the need to treat the active LRTI. Under the new therapeutic regimen both of the patient's chief complaints were addressed while reducing the overall risk for adverse effects due to DDIs.

Acute lower respiratory infections. European Lung white book. European Respiratory Society Accessed November 18, Cipro I.

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