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Cofactor specificity motifs and the induced fit mechanism in class I ketol-acid reductoisomerases

Cahn, Jackson K. B. and Brinkmann-Chen, Sabine and Spatzal, Thomas and Wiig, Jared A. and Buller, Andrew R. and Einsle, Oliver and Hu, Yilin and Ribbe, Markus W. and Arnold, Frances H. (2015) Cofactor specificity motifs and the induced fit mechanism in class I ketol-acid reductoisomerases. Biochemical Journal, 468 (3). pp. 475-484. ISSN 0264-6021. PMCID PMC4550535. https://resolver.caltech.edu/CaltechAUTHORS:20151001-082128409

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Abstract

Although most sequenced members of the industrially important ketol-acid reductoisomerase (KARI) family are class I enzymes, structural studies to date have focused primarily on the class II KARIs, which arose through domain duplication. In the present study, we present five new crystal structures of class I KARIs. These include the first structure of a KARI with a six-residue β2αB (cofactor specificity determining) loop and an NADPH phosphate-binding geometry distinct from that of the seven- and 12-residue loops. We also present the first structures of naturally occurring KARIs that utilize NADH as cofactor. These results show insertions in the specificity loops that confounded previous attempts to classify them according to loop length. Lastly, we explore the conformational changes that occur in class I KARIs upon binding of cofactor and metal ions. The class I KARI structures indicate that the active sites close upon binding NAD(P)H, similar to what is observed in the class II KARIs of rice and spinach and different from the opening of the active site observed in the class II KARI of Escherichia coli. This conformational change involves a decrease in the bending of the helix that runs between the domains and a rearrangement of the nicotinamide-binding site.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1042/BJ20150183DOIArticle
http://www.biochemj.org/content/468/3/475PublisherArticle
http://www.biochemj.org/content/468/3/475.supplementalPublisherSupplemental Material
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4550535PubMed CentralArticle
ORCID:
AuthorORCID
Brinkmann-Chen, Sabine0000-0002-5419-4192
Spatzal, Thomas0000-0002-9136-5915
Buller, Andrew R.0000-0002-9635-4844
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2015 The Authors Journal compilation © 2015 Biochemical Society. Received 11 February 2015/1 April 2015; accepted 7 April 2015. Published as BJ Immediate Publication 7 April 2015, doi:10.1042/BJ20150183 This work was supported by the Gordon and Betty Moore Foundation [grant number GBMF2809] to the Caltech Programmable Molecular Technology Initiative; the Resnick Sustainability Institute at Caltech (to J.K.B.C.); the National Institute of Health [grant number GM 67626 (to M.W.R.)]; the Molecular Observatory is supported by the Gordon and Betty Moore Foundation, the Beckman Institute, and the Sanofi-Aventis Bioengineering Research Program at Caltech. We thank Dr Jens Kaiser and Pavle Nikolovski for their continued support. AUTHOR CONTRIBUTION: Jackson Cahn and Sabine Brinkmann-Chen conceived of the project. Sabine Brinkmann-Chen prepared and crystallized three proteins, Jackson Cahn one and Jared Wiig and Thomas Spatzal one. Jackson Cahn solved the protein crystal structures, one in collaboration with Thomas Spatzal and one with Andrew Buller; Yilin Hu and Markus Ribbe provided guidance to Jared Wiig, Oliver Einsle to Thomas Spatzal and Frances Arnold to Jackson Cahn, Sabine Brinkmann-Chen and Andrew Buller. Sabine Brinkmann-Chen measured the thermostability of one of the KARIs; Jackson Cahn analysed the crystal structures and prepared the figures; Jackson Cahn, Sabine Brinkmann-Chen, Andrew Buller and Frances Arnold wrote the manuscript. Coordinates and structure factors have been deposited in the Protein Data Bank with accession numbers 4TSK, 4XDY, 4XDZ, 4XEH and 4XIY, as shown in Table 1.
Group:Resnick Sustainability Institute
Funders:
Funding AgencyGrant Number
Gordon and Betty Moore FoundationGBMF2809
Resnick Sustainability InstituteUNSPECIFIED
NIHGM 67626
Caltech Beckman InstituteUNSPECIFIED
Caltech Sanofi-Aventis Bioengineering Research ProgramUNSPECIFIED
Subject Keywords:acetohydroxyacid isomeroreductase (AHAIR), cofactor binding, conformational change, crystal structure, ketol-acid reductoisomerase (KARI)
Issue or Number:3
PubMed Central ID:PMC4550535
Record Number:CaltechAUTHORS:20151001-082128409
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20151001-082128409
Official Citation:Cofactor specificity motifs and the induced fit mechanism in class I ketol-acid reductoisomerases Jackson K.B. Cahn, Sabine Brinkmann-Chen, Thomas Spatzal, Jared A. Wiig, Andrew R. Buller, Oliver Einsle, Yilin Hu, Markus W. Ribbe, Frances H. Arnold Biochemical Journal Jun 2015, 468 (3) 475-484; DOI: 10.1042/BJ20150183
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:60646
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:01 Oct 2015 17:12
Last Modified:14 Oct 2019 18:08

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