6JD2

Crystal structure of Sulfolobus solfataricus ketol-acid reductoisomerase (Sso-KARI) in complex with Mg2+ at pH8.5

Summary for 6JD2

• Entry DOI: 10.2210/pdb6jd2/pdb
• Descriptor: Putative ketol-acid reductoisomerase 2, BETA-MERCAPTOETHANOL, MAGNESIUM ION, ... (4 entities in total)
• Functional Keywords: bi-specific, thermostable, reductoisomerase, magnesium-dependent, dodecamer, knotted protein, isomerase
• Biological source: Saccharolobus solfataricus (strain ATCC 35092 / DSM 1617 / JCM 11322 / P2) (Sulfolobus solfataricus)
• Total number of polymer chains: 12
• Total formula weight: 448279.18

Authors

Chen, C.Y.,Chang, Y.C.,Lin, K.F.,Huang, C.H.,Lin, B.L.,Ko, T.P.,Hsieh, D.L.,Tsai, M.D. (deposition date: 2019-01-30, release date: 2019-08-14, Last modification date: 2023-11-22)

Primary citation

Chen, C.Y.,Chang, Y.C.,Lin, B.L.,Lin, K.F.,Huang, C.H.,Hsieh, D.L.,Ko, T.P.,Tsai, M.D.
Use of Cryo-EM To Uncover Structural Bases of pH Effect and Cofactor Bispecificity of Ketol-Acid Reductoisomerase.
J.Am.Chem.Soc., 141:6136-6140, 2019

PubMed Abstract: While cryo-EM is revolutionizing structural biology, its impact on enzymology is yet to be fully demonstrated. The ketol-acid reductoisomerase (KARI) catalyzes conversion of (2 S)-acetolactate or (2 S)-aceto-2-hydroxybutyrate to 2,3-dihydroxy-3-alkylbutyrate. We found that KARI from archaea Sulfolobus solfataricus (Sso-KARI) is unusual in being a dodecamer, bispecific to NADH and NADPH, and losing activity above pH 7.8. While crystals were obtainable only at pH 8.5, cryo-EM structures were solved at pH 7.5 and 8.5 for Sso-KARI:2Mg. The results showed that the distances of the two catalytic Mg ions are lengthened in both structures at pH 8.5. We next solved cryo-EM structures of two Sso-KARI complexes, with NADH+inhibitor and NADPH+inhibitor at pH 7.5, which indicate that the bispecificity can be attributed to a unique asparagine at the cofactor binding loop. Unexpectedly, Sso-KARI also differs from other KARI enzymes in lacking "induced-fit", reflecting structural rigidity. Thus, cryo-EM is powerful for structural and mechanistic enzymology.

PubMed: 30921515
DOI: 10.1021/jacs.9b01354

Experimental method

X-RAY DIFFRACTION (2.53 Å)