Structural insight into the catalytic mechanism of gluconate 5-dehydrogenase fromStreptococcus suis: Crystal structures of the substrate-free and quaternary complex enzymes

The soil bacterium, Pseudomonas putida, is capable of using the alicyclic compound quinate as a sole carbon source. During this process, quinate is converted to 3-dehydroshikimate, which subsequently undergoes a dehydration to form protocatechuate. The latter transformation is performed by the enzyme dehydroshikimate dehydratase (DSD). We have recombinantly produced DSD from P. putida and are currently performing x-ray crystallographic studies on the enzyme to gain structural insight into its catalytic mechanism and mode of substrate recognition. Initial crystals of DSD diffracted to 2.7 Ä resolution, but exhibited strong twinning. A redesigned construct has recently yielded crystals that diffract to similar resolution, but with a significantly reduced tendency toward twinning. Interestingly, sequence analysis of P. putida DSD reveals that the protein is in fact a fusion of two distinct domains: an N-terminal sugar phosphate isomerase-like domain associated with DSD activity, and a C-terminal hydroxyphenylpyruvate dioxygenase (HPPD)-like domain with unknown functional significance. Structural characterization of the protein may provide novel insight into the functional relevance of the unusual HPPD-like domain.

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Structural insight into the catalytic mechanism of a cis-epoxysuccinate hydrolase producing enantiomerically pure d(−)-tartaric acid

Chemical Communications ◽

10.1039/c8cc04398a ◽

2018 ◽

Vol 54 (61) ◽

pp. 8482-8485 ◽

Cited By ~ 2

Author(s):

Sheng Dong ◽

Xi Liu ◽

Gu-Zhen Cui ◽

Qiu Cui ◽

Xinquan Wang ◽

...

Keyword(s):

Tartaric Acid ◽

Catalytic Mechanism ◽

Enantiomerically Pure ◽

Tartaric Acids ◽

Structural Insight ◽

Insight Into ◽

High Stereoselectivity

The catalytic mechanism for the high stereoselectivity and product enantioselectivity of a cis-epoxysuccinate hydrolase producing d(−)-tartaric acids was elucidated.

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Structures of AzrA and of AzrC complexed with substrate or inhibitor: insight into substrate specificity and catalytic mechanism

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s1399004713030988 ◽

2014 ◽

Vol 70 (2) ◽

pp. 553-564 ◽

Cited By ~ 14

Author(s):

Jian Yu ◽

Daiki Ogata ◽

ZuoQi Gai ◽

Seiichi Taguchi ◽

Isao Tanaka ◽

...

Keyword(s):

Substrate Specificity ◽

Crystal Structures ◽

Azo Dyes ◽

Catalytic Mechanism ◽

Initial Step ◽

Site Directed Mutagenesis ◽

Flavin Mononucleotide ◽

Catalytic Mechanisms ◽

Hydroxy Groups ◽

Insight Into

Azo dyes are major synthetic dyestuffs with one or more azo bonds and are widely used for various industrial purposes. The biodegradation of residual azo dyesviaazoreductase-catalyzed cleavage is very efficient as the initial step of wastewater treatment. The structures of the complexes of azoreductases with various substrates are therefore indispensable to understand their substrate specificity and catalytic mechanism. In this study, the crystal structures of AzrA and of AzrC complexed with Cibacron Blue (CB) and the azo dyes Acid Red 88 (AR88) and Orange I (OI) were determined. As an inhibitor/analogue of NAD(P)H, CB was located on top of flavin mononucleotide (FMN), suggesting a similar binding manner as NAD(P)H for direct hydride transfer to FMN. The structures of the AzrC–AR88 and AzrC–OI complexes showed two manners of binding for substrates possessing a hydroxy group at theorthoor theparaposition of the azo bond, respectively, while AR88 and OI were estimated to have a similar binding affinity to AzrC from ITC experiments. Although the two substrates were bound in different orientations, the hydroxy groups were located in similar positions, resulting in an arrangement of electrophilic C atoms binding with a proton/electron-donor distance of ∼3.5 Å to N5 of FMN. Catalytic mechanisms for different substrates are proposed based on the crystal structures and on site-directed mutagenesis analysis.

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Structural insight into poly(A) binding and catalytic mechanism of human PARN

The EMBO Journal ◽

10.1038/sj.emboj.7600869 ◽

2005 ◽

Vol 24 (23) ◽

pp. 4082-4093 ◽

Cited By ~ 78

Author(s):

Mousheng Wu ◽

Michael Reuter ◽

Hauke Lilie ◽

Yuying Liu ◽

Elmar Wahle ◽

...

Keyword(s):

Catalytic Mechanism ◽

Structural Insight ◽

Insight Into

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Structural insight into the catalytic mechanism of arsenate reductase from Synechocystis sp. PCC 6803

Environmental Arsenic in a Changing World ◽

10.1201/9781351046633-37 ◽

2019 ◽

pp. 95-96

Author(s):

Y. Yan ◽

J. Ye ◽

X. Zhang ◽

X.M. Xue ◽

Y.G. Zhu

Keyword(s):

Catalytic Mechanism ◽

Arsenate Reductase ◽

Structural Insight ◽

Insight Into ◽

Synechocystis Sp ◽

Pcc 6803

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Crystal Structures of Creatininase Reveal the Substrate Binding Site and Provide an Insight into the Catalytic Mechanism

Journal of Molecular Biology ◽

10.1016/j.jmb.2004.01.022 ◽

2004 ◽

Vol 337 (2) ◽

pp. 399-416 ◽

Cited By ~ 13

Author(s):

Tadashi Yoshimoto ◽

Nobutada Tanaka ◽

Naota Kanada ◽

Takahiko Inoue ◽

Yoshitaka Nakajima ◽

...

Keyword(s):

Binding Site ◽

Crystal Structures ◽

Catalytic Mechanism ◽

Substrate Binding ◽

Substrate Binding Site ◽

Insight Into

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Crystal structures of the CPAP/STIL complex reveal its role in centriole assembly and human microcephaly

eLife ◽

10.7554/elife.01071 ◽

2013 ◽

Vol 2 ◽

Cited By ~ 63

Author(s):

Matthew A Cottee ◽

Nadine Muschalik ◽

Yao Liang Wong ◽

Christopher M Johnson ◽

Steven Johnson ◽

...

Keyword(s):

Complex Formation ◽

Crystal Structures ◽

Brain Size ◽

Point Mutations ◽

Centriole Duplication ◽

Structural Insight ◽

Centriole Assembly ◽

Cilia And Flagella ◽

Insight Into

Centrioles organise centrosomes and template cilia and flagella. Several centriole and centrosome proteins have been linked to microcephaly (MCPH), a neuro-developmental disease associated with small brain size. CPAP (MCPH6) and STIL (MCPH7) are required for centriole assembly, but it is unclear how mutations in them lead to microcephaly. We show that the TCP domain of CPAP constitutes a novel proline recognition domain that forms a 1:1 complex with a short, highly conserved target motif in STIL. Crystal structures of this complex reveal an unusual, all-β structure adopted by the TCP domain and explain how a microcephaly mutation in CPAP compromises complex formation. Through point mutations, we demonstrate that complex formation is essential for centriole duplication in vivo. Our studies provide the first structural insight into how the malfunction of centriole proteins results in human disease and also reveal that the CPAP–STIL interaction constitutes a conserved key step in centriole biogenesis.

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