PDBsum entry 6acf

Oxidoreductase

PDB id: 6acf

Contents

Protein chains

(+ 2 more) 367 a.a.

PDB id:

6acf

Name:

Oxidoreductase

Title:

Structure of leucine dehydrogenase from geobacillus stearothermophilus by cryo-em

Structure:

Leucine dehydrogenase. Chain: a, b, c, d, e, f, g, h. Engineered: yes

Source:

Geobacillus stearothermophilus 10. Organism_taxid: 272567. Gene: gt50_15010. Expressed in: escherichia coli. Expression_system_taxid: 562

Authors:

H.Yamaguchi,A.Kamegawa,K.Nakata,T.Kashiwagi,T.Mizukoshi,Y.Fujiyoshi, K.Tani

Key ref:

H.Yamaguchi et al. (2019). Structural insights into thermostabilization of leucine dehydrogenase from its atomic structure by cryo-electron microscopy. J Struct Biol, 205, 11-21. PubMed id: 30543982 DOI: 10.1016/j.jsb.2018.12.001

Date:

26-Jul-18 Release date: 26-Dec-18

Protein chains

A0A0K2HC96  () -

DOI no: 10.1016/j.jsb.2018.12.001

J Struct Biol 205:11-21 (2019)

PubMed id: 30543982

Structural insights into thermostabilization of leucine dehydrogenase from its atomic structure by cryo-electron microscopy.

H.Yamaguchi, A.Kamegawa, K.Nakata, T.Kashiwagi, T.Mizukoshi, Y.Fujiyoshi, K.Tani.

ABSTRACT

Leucine dehydrogenase (LDH, EC 1.4.1.9) is a NAD⁺-dependent oxidoreductase that catalyzes the deamination of branched-chain l-amino acids (BCAAs). LDH of Geobacillus stearothermophilus (GstLDH) is a highly thermostable enzyme that has been applied for the quantification or production of BCAAs. Here the cryo-electron microscopy (cryo-EM) structures of apo and NAD⁺-bound LDH are reported at 3.0 and 3.2 Å resolution, respectively. On comparing the structures, the two overall structures are almost identical, but it was observed that the partial conformational change was triggered by the interaction between Ser147 and the nicotinamide moiety of NAD⁺. NAD⁺ binding also enhanced the strength of oligomerization interfaces formed by the core domains. Such additional interdomain interaction is in good agreement with our experimental results showing that the residual activity of NAD⁺-bound form was approximately three times higher than that of the apo form after incubation at 80 °C. In addition, sequence comparison of three structurally known LDHs indicated a set of candidates for site-directed mutagenesis to improve thermostability. Subsequent mutation analysis actually revealed that non-conserved residues, including Ala94, Tyr127, and the C-terminal region, are crucial for oligomeric thermostability.