Abstract
Nitrophorin 4 (NP4) belongs to a family of pH-sensitive, nitric oxide (NO) transporter proteins that undergo a large structural change from a closed to an open conformation at high pH to allow for NO delivery. Measuring the pH-dependent structural dynamics in NP4-NO around the ligand binding site is crucial for developing a mechanistic understanding of NO binding and release. In this study, we use coherent two-dimensional infrared (2D IR) spectroscopy to measure picosecond structural dynamics sampled by the nitrosyl stretch in NP4-NO as a function of pH at room temperature. Our results show that both the closed and open conformers of the protein are present at low (pD 5.1) and high (pD 7.9) pH conditions. The closed and open conformers are characterized by two frequencies of the nitrosyl stretching vibration labeled A0 and A1, respectively. Analysis of the 2D IR line shapes reveals that at pD 5.1, the closed conformer experiences structural fluctuations arising from solvation dynamics on a ∼3 ps time scale. At pD 7.9, both the open and closed conformers exhibit fluctuations on a ∼1 ps time scale. At both pD conditions, the closed conformers maintain a static distribution of structures within the experimental time window of 100 ps. This is in contrast to the open conformer, which is able to interconvert among its substates on a ∼100 ps time scale. Our results directly measure the time scales of solvation dynamics in the distal pocket, the flexibility of the open conformation at high pH, and the rigidity of the closed conformers at both pH conditions. We discuss how the pH-dependent equilibrium structural fluctuations of the nitrosyl ligand measured in this study are related to the uptake and delivery of nitric oxide in NP4.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 15804-15811 |
| Number of pages | 8 |
| Journal | Journal of Physical Chemistry B |
| Volume | 117 |
| Issue number | 49 |
| DOIs | |
| State | Published - Dec 12 2013 |
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ASJC Scopus subject areas
- Physical and Theoretical Chemistry
- Materials Chemistry
- Surfaces, Coatings and Films
Cite this
PH-dependent picosecond structural dynamics in the distal pocket of nitrophorin 4 investigated by 2D IR spectroscopy. / Cheng, Mark; Brookes, Jennifer F.; Montfort, William "Bill"; Khalil, Munira.
In: Journal of Physical Chemistry B, Vol. 117, No. 49, 12.12.2013, p. 15804-15811.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - PH-dependent picosecond structural dynamics in the distal pocket of nitrophorin 4 investigated by 2D IR spectroscopy
AU - Cheng, Mark
AU - Brookes, Jennifer F.
AU - Montfort, William "Bill"
AU - Khalil, Munira
PY - 2013/12/12
Y1 - 2013/12/12
N2 - Nitrophorin 4 (NP4) belongs to a family of pH-sensitive, nitric oxide (NO) transporter proteins that undergo a large structural change from a closed to an open conformation at high pH to allow for NO delivery. Measuring the pH-dependent structural dynamics in NP4-NO around the ligand binding site is crucial for developing a mechanistic understanding of NO binding and release. In this study, we use coherent two-dimensional infrared (2D IR) spectroscopy to measure picosecond structural dynamics sampled by the nitrosyl stretch in NP4-NO as a function of pH at room temperature. Our results show that both the closed and open conformers of the protein are present at low (pD 5.1) and high (pD 7.9) pH conditions. The closed and open conformers are characterized by two frequencies of the nitrosyl stretching vibration labeled A0 and A1, respectively. Analysis of the 2D IR line shapes reveals that at pD 5.1, the closed conformer experiences structural fluctuations arising from solvation dynamics on a ∼3 ps time scale. At pD 7.9, both the open and closed conformers exhibit fluctuations on a ∼1 ps time scale. At both pD conditions, the closed conformers maintain a static distribution of structures within the experimental time window of 100 ps. This is in contrast to the open conformer, which is able to interconvert among its substates on a ∼100 ps time scale. Our results directly measure the time scales of solvation dynamics in the distal pocket, the flexibility of the open conformation at high pH, and the rigidity of the closed conformers at both pH conditions. We discuss how the pH-dependent equilibrium structural fluctuations of the nitrosyl ligand measured in this study are related to the uptake and delivery of nitric oxide in NP4.
AB - Nitrophorin 4 (NP4) belongs to a family of pH-sensitive, nitric oxide (NO) transporter proteins that undergo a large structural change from a closed to an open conformation at high pH to allow for NO delivery. Measuring the pH-dependent structural dynamics in NP4-NO around the ligand binding site is crucial for developing a mechanistic understanding of NO binding and release. In this study, we use coherent two-dimensional infrared (2D IR) spectroscopy to measure picosecond structural dynamics sampled by the nitrosyl stretch in NP4-NO as a function of pH at room temperature. Our results show that both the closed and open conformers of the protein are present at low (pD 5.1) and high (pD 7.9) pH conditions. The closed and open conformers are characterized by two frequencies of the nitrosyl stretching vibration labeled A0 and A1, respectively. Analysis of the 2D IR line shapes reveals that at pD 5.1, the closed conformer experiences structural fluctuations arising from solvation dynamics on a ∼3 ps time scale. At pD 7.9, both the open and closed conformers exhibit fluctuations on a ∼1 ps time scale. At both pD conditions, the closed conformers maintain a static distribution of structures within the experimental time window of 100 ps. This is in contrast to the open conformer, which is able to interconvert among its substates on a ∼100 ps time scale. Our results directly measure the time scales of solvation dynamics in the distal pocket, the flexibility of the open conformation at high pH, and the rigidity of the closed conformers at both pH conditions. We discuss how the pH-dependent equilibrium structural fluctuations of the nitrosyl ligand measured in this study are related to the uptake and delivery of nitric oxide in NP4.
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U2 - 10.1021/jp407052a
DO - 10.1021/jp407052a
M3 - Article
C2 - 23885811
AN - SCOPUS:84887595106
VL - 117
SP - 15804
EP - 15811
JO - Journal of Physical Chemistry B Materials
JF - Journal of Physical Chemistry B Materials
SN - 1520-6106
IS - 49
ER -