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Crystallographic binding studies of rat peroxisomal multifunctional enzyme type 1 with 3‐ketodecanoyl‐CoA: capturing active and inactive states of its hydratase and dehydrogenase catalytic sites
Author(s) -
Sridhar Shruthi,
Schmitz Werner,
Hiltunen J. Kalervo,
Venkatesan Rajaram,
Bergmann Ulrich,
Kiema Tiila-Riikka,
Wierenga Rikkert K.
Publication year - 2020
Publication title -
acta crystallographica section d
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.374
H-Index - 138
ISSN - 2059-7983
DOI - 10.1107/s2059798320013819
Subject(s) - active site , linker , nad+ kinase , dehydrogenase , stereochemistry , helix (gastropod) , chemistry , peroxisome , enzyme , crystallography , biochemistry , biology , receptor , ecology , snail , computer science , operating system
The peroxisomal multifunctional enzyme type 1 (MFE1) catalyzes two successive reactions in the β‐oxidation cycle: the 2 E ‐enoyl‐CoA hydratase (ECH) and NAD + ‐dependent 3 S ‐hydroxyacyl‐CoA dehydrogenase (HAD) reactions. MFE1 is a monomeric enzyme that has five domains. The N‐terminal part (domains A and B) adopts the crotonase fold and the C‐terminal part (domains C, D and E) adopts the HAD fold. A new crystal form of MFE1 has captured a conformation in which both active sites are noncompetent. This structure, at 1.7 Å resolution, shows the importance of the interactions between Phe272 in domain B (the linker helix; helix H10 of the crotonase fold) and the beginning of loop 2 (of the crotonase fold) in stabilizing the competent ECH active‐site geometry. In addition, protein crystallographic binding studies using optimized crystal‐treatment protocols have captured a structure with both the 3‐ketodecanoyl‐CoA product and NAD + bound in the HAD active site, showing the interactions between 3‐ketodecanoyl‐CoA and residues of the C, D and E domains. Structural comparisons show the importance of domain movements, in particular of the C domain with respect to the D/E domains and of the A domain with respect to the HAD part. These comparisons suggest that the N‐terminal part of the linker helix, which interacts tightly with domains A and E, functions as a hinge region for movement of the A domain with respect to the HAD part.