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X‐Ray Crystal Structure of a Mutant Assimilatory Nitrite Reductase That Shows Sulfite Reductase‐Like Activity
Author(s) -
Nakano Shogo,
Takahashi Misa,
Sakamoto Atsushi,
Morikawa Hiromichi,
Katayanagi Katsuo
Publication year - 2012
Publication title -
chemistry and biodiversity
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.427
H-Index - 70
eISSN - 1612-1880
pISSN - 1612-1872
DOI - 10.1002/cbdv.201100442
Subject(s) - chemistry , sulfite reductase , sulfite , nitrite reductase , conformational change , reductase , ligand (biochemistry) , substrate (aquarium) , nitrite , stereochemistry , ammonium , mutant , crystal structure , nitrate reductase , crystallography , enzyme , biochemistry , nitrate , organic chemistry , oceanography , receptor , geology , gene
Assimilatory nitrite reductase (aNiR) reduces nitrite ions (NO $\rm{{_{2}^{-}}}$ ) to ammonium ions (NH $\rm{{_{4}^{+}}}$ ), whereas assimilatory sulfite reductase reduces sulfite (SO $\rm{{_{3}^{2-}}}$ ) to hydrogen sulfide (HS − ). Although aNiR can also reduce SO $\rm{{_{3}^{2-}}}$ , its activity is much lower than when NO $\rm{{_{2}^{-}}}$ is reduced as the substrate. To increase the SO $\rm{{_{3}^{2-}}}$ ‐reduction activity of aNiR, we performed a N226K mutation of Nii3, a representative aNiR. The resulting Nii3‐N226K variant could bind non‐native targets, SO $\rm{{_{3}^{2-}}}$ , and HCO $\rm{{_{3}^{-}}}$ , in addition to its native target, i.e. , NO $\rm{{_{2}^{-}}}$ . We have determined the high‐resolution structure of Nii3‐N226K in its apo‐state and in complex with SO $\rm{{_{3}^{2-}}}$ , NO $\rm{{_{2}^{-}}}$ , and HCO $\rm{{_{3}^{-}}}$ . This analysis revealed conformational changes of Lys226 and the adjacent Lys224 upon binding of SO $\rm{{_{3}^{2-}}}$ , but not NO $\rm{{_{2}^{-}}}$ . In contrast, HCO $\rm{{_{3}^{-}}}$ binding induced a conformational change at Arg179. After replacing Asn226 with a positively charged Lys, aNiR showed affinity for several anions. A comparison of all ligand‐bound structures for Nii3‐N226K revealed that structural changes in the active site depend on the size of the substrate.