Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Disruption of methylarginine metabolism impairs vascular homeostasis

Nature Medicine volume 13, pages 198–203 (2007)Cite this article

Abstract

Asymmetric dimethylarginine (ADMA) and monomethyl arginine (L-NMMA) are endogenously produced amino acids that inhibit all three isoforms of nitric oxide synthase (NOS)1. ADMA accumulates in various disease states, including renal failure, diabetes and pulmonary hypertension, and its concentration in plasma is strongly predictive of premature cardiovascular disease and death2,3,4. Both L-NMMA and ADMA are eliminated largely through active metabolism by dimethylarginine dimethylaminohydrolase (DDAH)5 and thus DDAH dysfunction may be a crucial unifying feature of increased cardiovascular risk. However, despite considerable interest in this pathway and in the role of ADMA as a cardiovascular risk factor, there is little evidence to support a causal role of ADMA in pathophysiology. Here we reveal the structure of human DDAH-1 and probe the function of DDAH-1 both by deleting the DDAH1 gene in mice and by using DDAH-specific inhibitors which, as we demonstrate by crystallography, bind to the active site of human DDAH-1. We show that loss of DDAH-1 activity leads to accumulation of ADMA and reduction in NO signaling. This in turn causes vascular pathophysiology, including endothelial dysfunction, increased systemic vascular resistance and elevated systemic and pulmonary blood pressure. Our results also suggest that DDAH inhibition could be harnessed therapeutically to reduce the vascular collapse associated with sepsis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Generation of DDAH-1-deficient mice and selective inhibitors of DDAH-1.
Figure 2: Functional characterization of vascular effects of gene deletion of Ddah1 and chemical inhibition of DDAH.
Figure 3: In vivo and pulmonary vascular characterization of effects of reduced DDAH activity.
Figure 4: Inhibition of DDAH reverses excess production of nitric oxide in endotoxic shock.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Vallance, P., Leone, A., Calver, A., Collier, J. & Moncada, S. Accumulation of an endogenous inhibitor of nitric oxide synthesis in chronic renal failure. Lancet 339, 572–575 (1992).

    Article  CAS  Google Scholar 

  2. Schnabel, R. et al. Asymmetric dimethylarginine and the risk of cardiovascular events and death in patients with coronary artery disease: results from the AtheroGene study. Circ. Res. 97, e53–e59 (2005).

    Article  CAS  Google Scholar 

  3. Ravani, P. et al. Asymmetrical dimethylarginine predicts progression to dialysis and death in patients with chronic kidney disease: a competing risks modeling approach. J. Am. Soc. Nephrol. 16, 2449–2455 (2005).

    Article  CAS  Google Scholar 

  4. Cooke, J.P. Asymmetric dimethylarginine: the uber marker? Circulation 109, 1813–1819 (2004).

    Article  Google Scholar 

  5. Vallance, P. & Leiper, J. Blocking NO synthesis: how, where and why? Nat. Rev. Drug Discov. 1, 939–950 (2002).

    Article  CAS  Google Scholar 

  6. McBride, A.E. & Silver, P.A. State of the arg: protein methylation at arginine comes of age. Cell 106, 5–8 (2001).

    Article  CAS  Google Scholar 

  7. Rossiter, S. et al. Selective substrate-based inhibitors of mammalian dimethylarginine dimethylaminohydrolase. J. Med. Chem. 48, 4670–4678 (2005).

    Article  CAS  Google Scholar 

  8. Murray-Rust, J. et al. Structural insights into the hydrolysis of cellular nitric oxide synthase inhibitors by dimethylarginine dimethylaminohydrolase. Nat. Struct. Biol. 8, 679–683 (2001).

    Article  CAS  Google Scholar 

  9. Leiper, J., Murray-Rust, J., McDonald, N. & Vallance, P. S-nitrosylation of dimethylarginine dimethylaminohydrolase regulates enzyme activity: further interactions between nitric oxide synthase and DDAH. Proc. Natl. Acad. Sci. USA 99, 13527–13532 (2002).

    Article  CAS  Google Scholar 

  10. Ichinose, F., Roberts, J.D. & Zapol, W.M. Inhaled nitric oxide a selective pulmonary vasodilator, current use and therapeutic potential. Circulation 109, 3106–3111 (2004).

    Article  Google Scholar 

  11. Pullamsetti, S. et al. Increased levels and reduced catabolism of asymmetric and symmetric dimethylarginine in pulmonary hypertension. FASEB J. 19, 1175–1177 (2005).

    Article  CAS  Google Scholar 

  12. Arrigoni, F.I., Vallance, P., Haworth, S.G. & Leiper, J.M. Metabolism of asymmetric dimethylarginines is regulated in the lung developmentally and with pulmonary hypertension induced by hypobaric hypoxia. Circulation 107, 1195–1201 (2003).

    Article  CAS  Google Scholar 

  13. Millatt, L.J. et al. Evidence for dysregulation of dimethylarginine dimethylaminohydrolase I in chronic hypxia-induced pulmonary hypertension. Circulation 108, 1493–1498 (2003).

    Article  CAS  Google Scholar 

  14. Smith, C.L., Anthony, S., Hubank, M., Leiper, J.M. & Vallance, P. Effects of ADMA upon gene expression: an insight into the pathophysiological significance of raised plasma ADMA. PLoS Med. 2, e264 (2005).

    Article  Google Scholar 

  15. Lane, K.B. et al. Heterozygous germline mutations in BMPR2, encoding a TGF beta receptor, cause familial primary pulmonary hypertension. Nat. Genet. 26, 81–84 (2000).

    Article  CAS  Google Scholar 

  16. Tran, C.T., Fox, M.F., Vallance, P. & Leiper, J.M. Chromosomal localization, gene structure, and expression of DDAH-1: comparison with DDAH-2 and implications for evolutionary origins. Genomics 68, 101–105 (2000).

    Article  CAS  Google Scholar 

  17. Collaborative Computational Project, Number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  18. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  Google Scholar 

  19. Teerlink, T., Nijveldt, R.J., de Jong, S. & Leeuwen, P.A. Determination of arginine, asymmetric dimethylarginine and symmetric dimethylarginine in human plasma and other biological samples by high-performance liquid chromatography. Anal. Biochem. 303, 131–137 (2002).

    Article  CAS  Google Scholar 

  20. Hollenberg, S.M. et al. Characterization of a hyperdynamic murine model of resuscitated sepsis using echocardiography. Am. J. Respir. Crit. Care Med. 164, 891–895 (2001).

    Article  CAS  Google Scholar 

  21. Nandi, M. et al. Pulmonary hypertension in a GTP-cyclohydrolase 1-deficient mouse. Circulation 111, 2086–2090 (2005).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank the following for experimental assistance: L. Newman, S. Jey, S. Thakker, H. Gill, V. Taylor and A. Cole. This work was funded by grants from the British Heart Foundation (PG20007 and PG/02/165/14797), the Wellcome Trust (065612/Z/01) and Medical Research Council (G0000002). P.V., J.L. and M.N. are part of the European Vascular Genomics Network funded by the European Union (contract number LSHM-CT-2003-503254). This study received financial support from the European Commission under the 6th Framework Programme (contract number LSHM-CT-2005-018725, PULMOTENSION). N.Q.M. and J.M.-R. are funded by Cancer Research UK.

Author information

Author notes

  1. Patrick Vallance

    Present address: Present address: Glaxo Smith Kline, Greenford Road, Greenford, Middlesex UB6 0HE, UK.,

Authors and Affiliations

  1. Division of Medicine, Centre for Clinical Pharmacology, University College London, London, WC1E 6JJ, UK

    James Leiper, Manasi Nandi, Belen Torondel, Mohammed Malaki, Sharon Rossiter, Shelagh Anthony, Melanie Madhani, Caroline Smith, Beata Wojciak-Stothard, Ray Stidwill & Patrick Vallance

  2. School of Crystallography, Birkbeck, Malet Street, London, WC1E 7HX, UK

    Judith Murray-Rust, Bernard O'Hara & Neil Q McDonald

  3. Structural Biology Laboratory, London Research Institute, Cancer Research UK, 44 Lincoln's Inn Fields, London, WC2A 3PX, UK

    Judith Murray-Rust & Neil Q McDonald

  4. Wolfson Institute for Biomedical Research, University College London, London, WC1E 6JJ, UK

    David Selwood & Alain Rudiger

Authors
  1. James Leiper
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Manasi Nandi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Belen Torondel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Judith Murray-Rust
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mohammed Malaki
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Bernard O'Hara
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sharon Rossiter
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Shelagh Anthony
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Melanie Madhani
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. David Selwood
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Caroline Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Beata Wojciak-Stothard
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Alain Rudiger
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Ray Stidwill
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Neil Q McDonald
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Patrick Vallance
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.L. generated the Ddah1 gene–deleted mice, oversaw all molecular and cellular studies and was involved in every aspect of the work reported in this paper. M.N. undertook experiments and oversaw all aspects of functional phenotyping and pharmacological experiments. M.N., R.S. and A.R. contributed to the in vivo cardiovascular experiments. M. Malaki., B.T., M.M. and S.A. contributed to the ex vivo cardiovascular experiments. C.S. helped characterize the molecular phenotype of the mice. B.W.-S. undertook experiments on pulmonary endothelial cells. S.R. designed and synthesied the DDAH inhibitors with D.S. B.O'H. expressed, purified and crystallised DDAH-1 and collected the X-ray data. J.M.-R. solved, built and refined all structures. N.Q.M. supervised the structural biology aspects of the project. P.V. was involved in formulating the hypothesis, and in the design, analysis and interpretation of all experiments. P.V., J.L., M.N., J.M.-R. and N.Q.M. wrote the final manuscript.

Corresponding author

Correspondence to Patrick Vallance.

Ethics declarations

Competing interests

UCL holds patents on DDAH and the chemical structures identified in this manuscript. The funding agencies (BHF and Wellcome Trust) and UCL and Birkbeck College stand to gain financially from these patents. Since this work was completed, P.V. left UCL and is now a full-time employee of GSK and holds shares in GSK. GSK has had no input into the work and has no financial interest in the work described in this paper.

Supplementary information

Supplementary Fig. 1

Construct development for DDAH-1 knockout mice and biochemical characterisation of DDAH1+/− mice. (PDF 170 kb)

Supplementary Fig. 2

Effect of DDAH inhibitors on NOS activity in vitro (PDF 22 kb)

Supplementary Fig. 3

Effect of L-291 on endothelium denuded rat aorta (PDF 10 kb)

Supplementary Fig. 4

Effects of i.v. administration of L-291 on mouse plasma levels. (PDF 11 kb)

Supplementary Fig. 5

Dimethylarginine levels and nitrite release from cultured primary pulmonary endothelial cells from mice. (PDF 14 kb)

Supplementary Fig. 6

Functional reactivity to calcium ionophore in isolated vessels from DDAH1+/− mice or mice treated with L-291 (PDF 25 kb)

Supplementary Fig. 7

Ex vivo analysis of pulmonary vasculature from DDAH1 +/− mice (PDF 313 kb)

Supplementary Fig. 8

Schematic summarising the main findings of study. (PDF 380 kb)

Supplementary Table 1

Crystallographic data collection and refinement (PDF 21 kb)

Supplementary Table 2

EC50 values from functional studies on isolated mouse aortic rings. (PDF 13 kb)

Supplementary Methods (PDF 47 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Leiper, J., Nandi, M., Torondel, B. et al. Disruption of methylarginine metabolism impairs vascular homeostasis. Nat Med 13, 198–203 (2007). https://doi.org/10.1038/nm1543

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm1543

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing