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. 2022 May 20;8(20):eabn2276.
doi: 10.1126/sciadv.abn2276. Epub 2022 May 20.

Revisiting the concept of peptide bond planarity in an iron-sulfur protein by neutron structure analysis

Yuya Hanazono  1   2 , Yu Hirano  2   3 , Kazuki Takeda  1 , Katsuhiro Kusaka  4 , Taro Tamada  2 , Kunio Miki  1
Affiliations

Revisiting the concept of peptide bond planarity in an iron-sulfur protein by neutron structure analysis

Yuya Hanazono et al. Sci Adv. .

Abstract

The planarity of the peptide bond is important for the stability and structure formation of proteins. However, substantial distortion of peptide bonds has been reported in several high-resolution structures and computational analyses. To investigate the peptide bond planarity, including hydrogen atoms, we report a 1.2-Å resolution neutron structure of the oxidized form of high-potential iron-sulfur protein. This high-resolution neutron structure shows that the nucleus positions of the amide protons deviate from the peptide plane and shift toward the acceptors. The planarity of the H─N─C═O plane depends strongly on the pyramidalization of the nitrogen atom. Moreover, the orientation of the amide proton of Cys75 is different in the reduced and oxidized states, possibly because of the electron storage capacity of the iron-sulfur cluster.

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Figures

Fig. 1.
Fig. 1.. Electron and nuclear density maps of oxidized HiPIP.
(A) Electron and nuclear density maps around the peptide bond between Pro12 and Thr13 and between Asn70 and Val71. The 2Fobs − Fcalc electron density map is shown in gray at a contour level of +4σ. The Fobs − Fcalc hydrogen omit nuclear density map is shown in green and orange at a contour level of +5σ and −5σ, respectively. (B) Difference between the center of electron density and nuclear density of the indole ring of Trp78. The Fobs − Fcalc hydrogen omit electron density map and Fobs − Fcalc hydrogen omit nuclear density map are shown in yellow (+4σ) and pink (−7.5σ), respectively. (C) Close-up view of Nε1─Hε1 and Cη2─Hη2 of Trp78. The Fobs − Fcalc hydrogen omit electron density map and Fobs − Fcalc hydrogen omit nuclear density map are shown in yellow (+4.5σ, +4σ, and +3.5σ) and pink (−8σ, −7.5σ, and −7σ), respectively.
Fig. 2.
Fig. 2.. Distortion of the peptide bond.
(A) Deviations of hydrogen atoms from the peptide plane. The Fobs − Fcalc hydrogen omit nuclear density map is shown in magenta. The amide hydrogen atoms of riding models are shown in green. The distance between amide hydrogen and oxygen atoms are shown in black. The angles of NH…O for experimentally determined models and riding models are shown in magenta and green, respectively. (B) Histogram of the frequency of differences in donor-hydrogen acceptor angles in single-acceptor hydrogen bonds between the experimentally refined model and the riding model. (C) Deviations of hydrogen atoms from the acceptor atoms in bifurcated hydrogen bonds. (D) Histogram of the frequency of N─H bond lengths for amide protons.
Fig. 3.
Fig. 3.. Position of hydrogen and oxygen atoms in the peptide bond.
(A) Histogram of the frequency of the deviation from the plane of the ω angle. The peptide bonds including the refined amide protons are counted. (B) Histogram of the frequency of the deviation from the plane of the dihedral angle of Hi─Ni─Ci-1═Oi-1 (ω′ angle). (C) Scatter plot of θC against θN. (D) Scatter plot of θN against ω′ angle. Linear regression is shown as a solid line. (E) Scatter plot of θC against the ω′ angle. (F) Side-view schematic of the peptide bond. The attraction of the amide proton by the acceptor atom shifts the Hi─Ni─Ci-1═Oi-1 plane.
Fig. 4.
Fig. 4.. Iron-sulfur cluster including hydrogen atoms.
(A) Hydrogen bond with Sγ of cysteine residues covalently bound to Fe atoms. N─H bond lengths and H…O distances are shown in red and black, respectively. (B) Position of the amide proton of Cys75 in the oxidized state. The distance between the amide proton of Cys75 and S3 atom is shown in black. The deviations from the Ci-1─Ni─Cαi plane of the amide proton of Cys75 and oxygen of Trp74 are shown in blue and red, respectively. (C) Positions of the amide proton of Cys75 and oxygen of Trp74 in the reduced state [Protein Data Bank identifier (PDB ID) 5D8V]. (D) Structure of the HiPIP–LH1-RC complex around the contact surface between HiPIP and RC (PDB ID 7C52). The structural changes of the amide proton of Cys75 and the oxygen of Trp74 between the oxidized and reduced states are indicated by arrows.
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