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Structural and functional features of the NAD(P) dependent Gfo/Idh/MocA protein family oxidoreductases.

ABSTRACT: The Gfo/Idh/MocA protein family contains a number of different proteins, which almost exclusively consist of NAD(P)-dependent oxidoreductases that have a diverse set of substrates, typically pyranoses. In this study, to clarify common structural features that would contribute to their function, the available crystal structures of the members of this family have been analyzed. Despite a very low sequence identity, the central features of the three-dimensional structures of the proteins are surprisingly similar. The members of the protein family have a two-domain structure consisting of a N-terminal nucleotide-binding domain and a C-terminal ?/?-domain. The C-terminal domain contributes to the substrate binding and catalysis, and contains a ??-motif with a central ?-helix carrying common essential amino acid residues. The ?-sheet of the ?/?-domain contributes to the oligomerization in most of the proteins in the family.

SUBMITTER: Taberman H

PROVIDER: S-EPMC4941211 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature

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Structural and functional features of the NAD(P) dependent Gfo/Idh/MocA protein family oxidoreductases.

Taberman Helena H Parkkinen Tarja T Rouvinen Juha J

Protein science : a publication of the Protein Society 20160201 4

The Gfo/Idh/MocA protein family contains a number of different proteins, which almost exclusively consist of NAD(P)-dependent oxidoreductases that have a diverse set of substrates, typically pyranoses. In this study, to clarify common structural features that would contribute to their function, the available crystal structures of the members of this family have been analyzed. Despite a very low sequence identity, the central features of the three-dimensional structures of the proteins are surpri ...[more]

PMID: 26749496

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Project description:BackgroundThe chicken avidin gene family consists of avidin and several avidin related genes (AVRs). Of these gene products, avidin is the best characterized and is known for its extremely high affinity for D-biotin, a property that is utilized in numerous modern life science applications. Recently, the AVR genes have been expressed as recombinant proteins, which have shown different biotin-binding properties as compared to avidin.ResultsIn the present study, we have employed multiple biochemical methods to better understand the structure-function relationship of AVR proteins focusing on AVR2. Firstly, we have solved the high-resolution crystal structure of AVR2 in complex with a bound ligand, D-biotin. The AVR2 structure reveals an overall fold similar to the previously determined structures of avidin and AVR4. Major differences are seen, especially at the 1-3 subunit interface, which is stabilized mainly by polar interactions in the case of AVR2 but by hydrophobic interactions in the case of AVR4 and avidin, and in the vicinity of the biotin binding pocket. Secondly, mutagenesis, competitive dissociation analysis and differential scanning calorimetry were used to compare and study the biotin-binding properties as well as the thermal stability of AVRs and avidin. These analyses pinpointed the importance of residue 109 for biotin binding and stability of AVRs. The I109K mutation increased the biotin-binding affinity of AVR2, whereas the K109I mutation decreased the biotin-binding affinity of AVR4. Furthermore, the thermal stability of AVR2(I109K) increased in comparison to the wild-type protein and the K109I mutation led to a decrease in the thermal stability of AVR4.ConclusionAltogether, this study broadens our understanding of the structural features determining the ligand-binding affinities and stability as well as the molecular evolution within the protein family. This novel information can be applied to further develop and improve the tools already widely used in avidin-biotin technology.

Dataset Information

Structural and functional features of the NAD(P) dependent Gfo/Idh/MocA protein family oxidoreductases.

Publications

Structural and functional features of the NAD(P) dependent Gfo/Idh/MocA protein family oxidoreductases.

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