Dimerization and enzymatic activity of fungal 17β-hydroxysteroid dehydrogenase from the short-chain dehydrogenase/reductase superfamily

Kristan, Katja; Deluca, Dominga; Adamski, Jerzy; Stojan, Jure; Rižner, Tea Lanišnik

doi:10.1186/1471-2091-6-28

Cited by 16 publications

(20 citation statements)

References 40 publications

(46 reference statements)

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“…Previously, effects of mutants that break the dimer at the A-C type interface have been investigated in 17b-HSDcl (Kristan et al 2005). Like most dimeric members of the SDR family, the functional form of 17b-HSDcl is an A-C type dimer; breaking this dimer by point mutations results in inactive monomers.…”

Section: Tetramerization Is Not Required For Scr Activitymentioning

confidence: 99%

“…In the tetrameric form, the subunits are always observed to assume 2-2-2 symmetry in which subunit interfaces are described by three orthogonal dyad axes, conventionally termed P, Q, and R (Tanaka et al 2001). On the other hand, dimeric SDR proteins are found to mimic the dimer interface of either the P-or the Q-symmetry-related interface in the tetramer (Grimm et al 2000;Kristan et al 2005). All available three-dimensional (3D) structures of SDR subunits display a characteristic Rossmann-fold structure consisting of a central seven-strand parallel b-sheet flanked by ahelices (Liese et al 1996;Oppermann et al 2003).…”

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confidence: 99%

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Crystal structure of a carbonyl reductase from Candida parapsilosis with anti‐Prelog stereospecificity

et al. 2008

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A novel short-chain (S)-1-phenyl-1,2-ethanediol dehydrogenase (SCR) from Candida parapsilosis exhibits coenzyme specificity for NADPH over NADH. It catalyzes an anti-Prelog type reaction to reduce 2-hydroxyacetophenone into (S)-1-phenyl-1,2-ethanediol. The coding gene was overexpressed in Escherichia coli and the purified protein was crystallized. The crystal structure of the apo-form was solved to 2.7 Å resolution. This protein forms a homo-tetramer with a broken 2-2-2 symmetry. The overall fold of each SCR subunit is similar to that of the known structures of other homologous alcohol dehydrogenases, although the latter usually form tetramers with perfect 2-2-2 symmetries. Additionally, in the apo-SCR structure, the entrance of the NADPH pocket is blocked by a surface loop. In order to understand the structure-function relationship of SCR, we carried out a number of mutagenesisenzymatic analyses based on the new structural information. First, mutations of the putative catalytic Ser-Tyr-Lys triad confirmed their functional role. Second, truncation of an N-terminal 31-residue peptide indicated its role in oligomerization, but not in catalytic activity. Similarly, a V270D point mutation rendered the SCR as a dimer, rather than a tetramer, without affecting the enzymatic activity. Moreover, the S67D/H68D double-point mutation inside the coenzyme-binding pocket resulted in a nearly 10-fold increase and a 20-fold decrease in the k cat /K M value when NADH and NADPH were used as cofactors, respectively, with k cat remaining essentially the same. This latter result provides a new example of a protein engineering approach to modify the coenzyme specificity in SCR and short-chain dehydrogenases/reductases in general.

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Section: Tetramerization Is Not Required For Scr Activitymentioning

confidence: 99%

mentioning

confidence: 99%

Crystal structure of a carbonyl reductase from Candida parapsilosis with anti‐Prelog stereospecificity

et al. 2008

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“…The importance of dimerization for the activity of SDRs offers a specific target for drugs to regulate the activity of any given SDR. 25 …”

Section: Discussionmentioning

confidence: 99%

“…22–24 Our findings also support the proposal that enzymatic activity relies on the dimerization of the protein as observed for other SDRs. 25 A model of functionally related RDH12 allowed prediction of the impact of disease-causing mutations within the catalytic domain, including the dinucleotide-binding site of the protein or the dimerization interface.…”

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confidence: 99%

Structural Insights into the Drosophila melanogaster Retinol Dehydrogenase, a Member of the Short-Chain Dehydrogenase/Reductase Family

et al. 2016

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The 11-cis-retinylidene chromophore of visual pigments isomerizes upon interaction with a photon, initiating a downstream cascade of signaling events that ultimately lead to visual perception. 11-cis-Retinylidene is regenerated through enzymatic transformations collectively called the visual cycle. The first and rate-limiting enzymatic reaction within this cycle, i.e., the reduction of all-trans-retinal to all-trans-retinol, is catalyzed by retinol dehydrogenases. Here, we determined the structure of Drosophila melanogaster photoreceptor retinol dehydrogenase (PDH) isoform C that belongs to the short-chain dehydrogenase/reductase (SDR) family. This is the first reported structure of a SDR that possesses this biologically important activity. Two crystal structures of the same enzyme grown under different conditions revealed a novel conformational change of the NAD+ cofactor, likely representing a change during catalysis. Amide hydrogen–deuterium exchange of PDH demonstrated changes in the structure of the enzyme upon dinucleotide binding. In D. melanogaster, loss of PDH activity leads to photoreceptor degeneration that can be partially rescued by transgenic expression of human RDH12. Based on the structure of PDH, we analyzed mutations causing Leber congenital amaurosis 13 in a homology model of human RDH12 to obtain insights into the molecular basis of RDH12 disease-causing mutations.

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“…The Dictyostelium discoideum protein DDB_G0291732 consists of 308 amino acids and is clearly related to the NmrA-like SDR superfamily; however, it exists as a monomer in solution (Fig. 1b), in contrast to common SDR-type enzymes, which are found to be either homodimers or hometetramers (Kristan et al, 2005;Oppermann et al, 2003). Primary-sequence analysis shows that it lacks the extreme N-terminal cofactor-binding Gly-rich region and that the active-site Tyr residue is changed to a His residue, suggesting that it may not have dehydrogenase/reductase activity but may play other physiological role(s) similar to those of the dinucleotide-sensing NmrA or HSCARG (Stammers et al, 2001;Zheng et al, 2007).…”

Section: Introductionmentioning

confidence: 99%

Crystallization and preliminary X-ray crystallographic analysis of the short-chain dehydrogenase/reductase-type DDB_G0291732 protein fromDictyostelium discoideum

2010

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The DDB_G0291732 gene product from Dictyostelium discoideum, which is a NmrA-like protein that belongs to the short-chain dehydrogenase/reductase superfamily but shows deviations in conserved sequence regions, has been crystallized by the hanging-drop vapour-diffusion method at 295 K. A 1.65 Å resolution data set was collected using synchrotron radiation. The crystals of DDB_G0291732 protein belonged to space group P2(1), with unit-cell parameters a=38.5, b=63.7, c=56.0 Å, β=91.7°. Assuming the presence of one molecule in the asymmetric unit, the solvent content was estimated to be about 38.1%.

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Dimerization and enzymatic activity of fungal 17β-hydroxysteroid dehydrogenase from the short-chain dehydrogenase/reductase superfamily

Abstract: Background: 17β-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17β-HSDcl) is a member of the short-chain dehydrogenase/reductase (SDR) superfamily. SDR proteins usually function as dimers or tetramers and 17β-HSDcl is also a homodimer under native conditions.

Cited by 16 publications

References 40 publications

Crystal structure of a carbonyl reductase from Candida parapsilosis with anti‐Prelog stereospecificity

Crystal structure of a carbonyl reductase from Candida parapsilosis with anti‐Prelog stereospecificity

Structural Insights into the Drosophila melanogaster Retinol Dehydrogenase, a Member of the Short-Chain Dehydrogenase/Reductase Family

Crystallization and preliminary X-ray crystallographic analysis of the short-chain dehydrogenase/reductase-type DDB_G0291732 protein fromDictyostelium discoideum

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