RT Journal Article
SR Electronic
T1 Sub-3 Å resolution structure of 110 kDa nitrite reductase determined by 200 kV cryogenic electron microscopy
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.07.12.199695
DO 10.1101/2020.07.12.199695
A1 Naruhiko Adachi
A1 Takahide Yamaguchi
A1 Toshio Moriya
A1 Masato Kawasaki
A1 Kotaro Koiwai
A1 Akira Shinoda
A1 Yusuke Yamada
A1 Fumiaki Yumoto
A1 Takamitsu Kohzuma
A1 Toshiya Senda
YR 2020
UL http://biorxiv.org/content/early/2020/07/12/2020.07.12.199695.abstract
AB Cu-containing nitrite reductases (NiRs), a well-studied family of 110 kDa enzymes, play central roles in denitrification and have over 100 Protein Data Bank entries. However, such issues as crystal packing, photoreduction, and lack of high pH cases have impeded structural analysis of the catalytic mechanism. Here we show the cryogenic electron microscopy (cryo-EM) structures of Achromobacter cycloclastes NiR (AcNiR) at 2.99 and 2.85 Å resolution with pH 6.2 and 8.1, respectively. Comprehensive comparisons with cryo-EM and 56 AcNiR crystal structures suggested crystallographic artifacts in residues 185–215 and His255 due to packing and photoreduction, respectively. With electron paramagnetic resonance spectroscopy, a newly developed map comparison method supported local structural changes at pH 8.1 around the type-2 Cu site, including His255 deprotonation. While the theoretical coordination error estimation of cryo-EM structures remains difficult, combined analysis using X-ray and cryo-EM structures will allow deeper insight into the local structural changes of proteins.Competing Interest StatementThe authors have declared no competing interest.