Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase

Mack L. Flinspach; Huiying Li; Joumana Jamal; Weiping Yang; Hui Huang; Jung Mi Hah; José Antonio Gómez-Vidal; Elizabeth A. Litzinger; Richard B. Silverman; Thomas L. Poulos

doi:10.1038/nsmb704

Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase

Mack L. Flinspach, Huiying Li, Joumana Jamal, Weiping Yang, Hui Huang, Jung Mi Hah, José Antonio Gómez-Vidal, Elizabeth A. Litzinger, Richard B. Silverman, Thomas L. Poulos^*

^*Corresponding author for this work

Chemistry

Research output: Contribution to journal › Article › peer-review

75 Scopus citations

Abstract

Three nitric oxide synthase (NOS) isoforms, eNOS, nNOS and iNOS, generate nitric oxide (NO) crucial to the cardiovascular, nervous and host defense systems, respectively. Development of isoform-selective NOS inhibitors is of considerable therapeutic importance. Crystal structures of nNOS-selective dipeptide inhibitors in complex with both nNOS and eNOS were solved and the inhibitors were found to adopt a curled conformation in nNOS but an extended conformation in eNOS. We hypothesized that a single-residue difference in the active site, Asp597 (nNOS) versus Asn368 (eNOS), is responsible for the favored binding in nNOS. In the D597N nNOS mutant crystal structure, a bound inhibitor switches to the extended conformation and its inhibition of nNOS decreases >200-fold. Therefore, a single-residue difference is responsible for more than two orders of magnitude selectivity in inhibition of nNOS over eNOS by L-N^ω-nitroarginine-containing dipeptide inhibitors.

Original language	English (US)
Pages (from-to)	54-59
Number of pages	6
Journal	Nature Structural and Molecular Biology
Volume	11
Issue number	1
DOIs	https://doi.org/10.1038/nsmb704
State	Published - Jan 2004

Funding

We thank the beamline staff at SSRL and ALS for their assistance during synchrotron data collection. This research was supported by US National Institutes of Health grants GM57353 (T.L.P) and GM49725 (R.B.S.). J.A.G. acknowledges NATO and the Spanish Ministry of Science and Technology for support.

ASJC Scopus subject areas

Molecular Biology
Structural Biology

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10.1038/nsmb704

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title = "Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase",

abstract = "Three nitric oxide synthase (NOS) isoforms, eNOS, nNOS and iNOS, generate nitric oxide (NO) crucial to the cardiovascular, nervous and host defense systems, respectively. Development of isoform-selective NOS inhibitors is of considerable therapeutic importance. Crystal structures of nNOS-selective dipeptide inhibitors in complex with both nNOS and eNOS were solved and the inhibitors were found to adopt a curled conformation in nNOS but an extended conformation in eNOS. We hypothesized that a single-residue difference in the active site, Asp597 (nNOS) versus Asn368 (eNOS), is responsible for the favored binding in nNOS. In the D597N nNOS mutant crystal structure, a bound inhibitor switches to the extended conformation and its inhibition of nNOS decreases >200-fold. Therefore, a single-residue difference is responsible for more than two orders of magnitude selectivity in inhibition of nNOS over eNOS by L-Nω-nitroarginine-containing dipeptide inhibitors.",

author = "Flinspach, {Mack L.} and Huiying Li and Joumana Jamal and Weiping Yang and Hui Huang and Hah, {Jung Mi} and G{\'o}mez-Vidal, {Jos{\'e} Antonio} and Litzinger, {Elizabeth A.} and Silverman, {Richard B.} and Poulos, {Thomas L.}",

note = "Funding Information: We thank the beamline staff at SSRL and ALS for their assistance during synchrotron data collection. This research was supported by US National Institutes of Health grants GM57353 (T.L.P) and GM49725 (R.B.S.). J.A.G. acknowledges NATO and the Spanish Ministry of Science and Technology for support.",

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AU - Flinspach, Mack L.

AU - Li, Huiying

AU - Jamal, Joumana

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AU - Huang, Hui

AU - Hah, Jung Mi

AU - Gómez-Vidal, José Antonio

AU - Litzinger, Elizabeth A.

AU - Silverman, Richard B.

AU - Poulos, Thomas L.

N1 - Funding Information: We thank the beamline staff at SSRL and ALS for their assistance during synchrotron data collection. This research was supported by US National Institutes of Health grants GM57353 (T.L.P) and GM49725 (R.B.S.). J.A.G. acknowledges NATO and the Spanish Ministry of Science and Technology for support.

PY - 2004/1

Y1 - 2004/1

N2 - Three nitric oxide synthase (NOS) isoforms, eNOS, nNOS and iNOS, generate nitric oxide (NO) crucial to the cardiovascular, nervous and host defense systems, respectively. Development of isoform-selective NOS inhibitors is of considerable therapeutic importance. Crystal structures of nNOS-selective dipeptide inhibitors in complex with both nNOS and eNOS were solved and the inhibitors were found to adopt a curled conformation in nNOS but an extended conformation in eNOS. We hypothesized that a single-residue difference in the active site, Asp597 (nNOS) versus Asn368 (eNOS), is responsible for the favored binding in nNOS. In the D597N nNOS mutant crystal structure, a bound inhibitor switches to the extended conformation and its inhibition of nNOS decreases >200-fold. Therefore, a single-residue difference is responsible for more than two orders of magnitude selectivity in inhibition of nNOS over eNOS by L-Nω-nitroarginine-containing dipeptide inhibitors.

AB - Three nitric oxide synthase (NOS) isoforms, eNOS, nNOS and iNOS, generate nitric oxide (NO) crucial to the cardiovascular, nervous and host defense systems, respectively. Development of isoform-selective NOS inhibitors is of considerable therapeutic importance. Crystal structures of nNOS-selective dipeptide inhibitors in complex with both nNOS and eNOS were solved and the inhibitors were found to adopt a curled conformation in nNOS but an extended conformation in eNOS. We hypothesized that a single-residue difference in the active site, Asp597 (nNOS) versus Asn368 (eNOS), is responsible for the favored binding in nNOS. In the D597N nNOS mutant crystal structure, a bound inhibitor switches to the extended conformation and its inhibition of nNOS decreases >200-fold. Therefore, a single-residue difference is responsible for more than two orders of magnitude selectivity in inhibition of nNOS over eNOS by L-Nω-nitroarginine-containing dipeptide inhibitors.

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Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase

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Rat neuronal NOS D597N mutant heme domain with L-N(omega)-nitroarginine-2,4-L-diaminobutyric amide bound

Bovine eNOS N368D single mutant with L-N(omega)-Nitroarginine-(4R)-Amino-L-Proline Amide Bound

Rat nNOS D597N mutant with L-N(omega)-Nitroarginine-(4R)-amino-L-proline amide bound

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Structural basis for dipeptide amide isoform-selective inhibition of neuronal nitric oxide synthase

Abstract

Funding

ASJC Scopus subject areas

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Datasets

Rat neuronal NOS D597N mutant heme domain with L-N(omega)-nitroarginine-2,4-L-diaminobutyric amide bound

Bovine eNOS N368D single mutant with L-N(omega)-Nitroarginine-(4R)-Amino-L-Proline Amide Bound

Rat nNOS D597N mutant with L-N(omega)-Nitroarginine-(4R)-amino-L-proline amide bound

Cite this