A 2.3 A resolution structure of chymosin complexed with a reduced bond inhibitor shows that the active site beta-hairpin flap is rearranged when compared with the native crystal structure

The crystal structure of a copper amine oxidase fromArthrobacter globiformiswas determined at 1.08 Å resolution with the use of low-molecular-weight polyethylene glycol (LMW PEG; average molecular weight ∼200) as a cryoprotectant. The final crystallographicRfactor andRfreewere 13.0 and 15.0%, respectively. Several molecules of LMW PEG were found to occupy cavities in the protein interior, including the active site, which resulted in a marked reduction in the overallBfactor and consequently led to a subatomic resolution structure for a relatively large protein with a monomer molecular weight of ∼70 000. About 40% of the presumed H atoms were observed as clear electron densities in theFo−Fcdifference map. Multiple minor conformers were also identified for many residues. Anisotropic displacement fluctuations were evaluated in the active site, which contains a post-translationally derived quinone cofactor and a Cu atom. Furthermore, diatomic molecules, most likely to be molecular oxygen, are bound to the protein, one of which is located in a region that had previously been proposed as an entry route for the dioxygen substrate from the central cavity of the dimer interface to the active site.

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High-resolution structure of an atypical α-phosphoglucomutase related to eukaryotic phosphomannomutases

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s0907444913017046 ◽

2013 ◽

Vol 69 (10) ◽

pp. 2008-2016 ◽

Cited By ~ 3

Author(s):

Przemyslaw Nogly ◽

Pedro M. Matias ◽

Matteo de Rosa ◽

Rute Castro ◽

Helena Santos ◽

...

Keyword(s):

Crystal Structure ◽

High Resolution ◽

Active Site ◽

Catalytic Mechanism ◽

Haloacid Dehalogenase ◽

Enzyme Active Site ◽

High Resolution Structure ◽

Dimeric Form ◽

Strict Specificity ◽

Resolution Structure

The first structure of a bacterial α-phosphoglucomutase with an overall fold similar to eukaryotic phosphomannomutases is reported. Unlike most α-phosphoglucomutases within the α-D-phosphohexomutase superfamily, it belongs to subclass IIb of the haloacid dehalogenase superfamily (HADSF). It catalyzes the reversible conversion of α-glucose 1-phosphate to glucose 6-phosphate. The crystal structure of α-phosphoglucomutase fromLactococcus lactis(APGM) was determined at 1.5 Å resolution and contains a sulfate and a glycerol bound at the enzyme active site that partially mimic the substrate. A dimeric form of APGM is present in the crystal and in solution, an arrangement that may be functionally relevant. The catalytic mechanism of APGM and its strict specificity towards α-glucose 1-phosphate are discussed.

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Crystal structure of chorismate mutase fromBurkholderia phymatum

Acta Crystallographica Section F Structural Biology Communications ◽

10.1107/s2053230x18002868 ◽

2018 ◽

Vol 74 (4) ◽

pp. 187-192 ◽

Cited By ~ 1

Author(s):

Oluwatoyin A. Asojo ◽

Sandhya Subramanian ◽

Jan Abendroth ◽

Ilyssa Exley ◽

Donald D. Lorimer ◽

...

Keyword(s):

Crystal Structure ◽

Drug Development ◽

Structural Genomics ◽

Active Site ◽

Chorismate Mutase ◽

Biological Warfare ◽

Beneficial Bacteria ◽

Sequence Identity ◽

Resolution Structure

The bacteriumBurkholderia phymatumis a promiscuous symbiotic nitrogen-fixating bacterium that belongs to one of the largest groups of Betaproteobacteria. OtherBurkholderiaspecies are known to cause disease in plants and animals, and some are potential agents for biological warfare. Structural genomics efforts include characterizing the structures of enzymes from pathways that can be targeted for drug development. As part of these efforts, chorismate mutase fromB. phymatumwas produced and crystallized, and a 1.95 Å resolution structure is reported. This enzyme shares less than 33% sequence identity with other homologs of known structure. There are two classes of chorismate mutase: AroQ and AroH. The bacterial subclass AroQγ has reported roles in virulence. Chorismate mutase fromB. phymatumhas the prototypical AroQγ topology and retains the characteristic chorismate mutase active site. This suggests that substrate-based chorismate mutase inhibitors will not be specific and are likely to affect beneficial bacteria such asB. phymatum.

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The Aspergillus fumigatus Sialidase Is a 3-Deoxy-d-glycero-d-galacto-2-nonulosonic Acid Hydrolase (KDNase)

Journal of Biological Chemistry ◽

10.1074/jbc.m110.207043 ◽

2011 ◽

Vol 286 (12) ◽

pp. 10783-10792 ◽

Cited By ~ 13

Author(s):

Judith C. Telford ◽

Juliana H. F. Yeung ◽

Guogang Xu ◽

Milton J. Kiefel ◽

Andrew G. Watts ◽

...

Keyword(s):

Crystal Structure ◽

Aspergillus Fumigatus ◽

Active Site ◽

Catalytic Mechanism ◽

Chair Conformation ◽

Boat Conformation ◽

Transition State Analog ◽

Acetylneuraminic Acid ◽

Nonulosonic Acid ◽

Resolution Structure

Aspergillus fumigatus is a filamentous fungus that can cause severe respiratory disease in immunocompromised individuals. A putative sialidase from A. fumigatus was recently cloned and shown to be relatively poor in cleaving N-acetylneuraminic acid (Neu5Ac) in comparison with bacterial sialidases. Here we present the first crystal structure of a fungal sialidase. When the apo structure was compared with bacterial sialidase structures, the active site of the Aspergillus enzyme suggested that Neu5Ac would be a poor substrate because of a smaller pocket that normally accommodates the acetamido group of Neu5Ac in sialidases. A sialic acid with a hydroxyl in place of an acetamido group is 2-keto-3-deoxynononic acid (KDN). We show that KDN is the preferred substrate for the A. fumigatus sialidase and that A. fumigatus can utilize KDN as a sole carbon source. A 1.45-Å resolution crystal structure of the enzyme in complex with KDN reveals KDN in the active site in a boat conformation and nearby a second binding site occupied by KDN in a chair conformation, suggesting that polyKDN may be a natural substrate. The enzyme is not inhibited by the sialidase transition state analog 2-deoxy-2,3-dehydro-N-acetylneuraminic acid (Neu5Ac2en) but is inhibited by the related 2,3-didehydro-2,3-dideoxy-d-glycero-d-galacto-nonulosonic acid that we show bound to the enzyme in a 1.84-Å resolution crystal structure. Using a fluorinated KDN substrate, we present a 1.5-Å resolution structure of a covalently bound catalytic intermediate. The A. fumigatus sialidase is therefore a KDNase with a similar catalytic mechanism to Neu5Ac exosialidases, and this study represents the first structure of a KDNase.

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Structural elucidation of the heterodimeric cis-prenyltransferase NgBR/DHDDS complex reveals novel insights in regulation of protein glycosylation

10.1101/2020.06.03.132209 ◽

2020 ◽

Author(s):

Ban Edani ◽

Kariona A. Grabińska ◽

Rong Zhang ◽

Eon Joo Park ◽

Benjamin Siciliano ◽

...

Keyword(s):

Crystal Structure ◽

Active Site ◽

Protein Glycosylation ◽

Rate Limiting Step ◽

Limiting Step ◽

Chain Synthesis ◽

Rate Limiting ◽

Activity Comparison ◽

Long Chain ◽

Resolution Structure

SummaryCis-prenyltransferase (cis-PTase) catalyzes the rate-limiting step in the synthesis of glycosyl carrier lipids required for protein glycosylation in the lumen of endoplasmic reticulum. Here we report the crystal structure of the human NgBR/DHDDS complex, which represents the first atomic resolution structure for any heterodimeric cis-PTase. The crystal structure sheds light on how NgBR stabilizes DHDDS through dimerization, participates in the enzyme’s active site through its C-terminal -RXG- motif, and how phospholipids markedly stimulate cis-PTase activity. Comparison of NgBR/DHDDS with homodimeric cis-PTase structures leads to a model where the elongating isoprene chain extends beyond the enzyme’s active site tunnel, and an insert within the α3 helix helps to stabilize this energetically unfavorable state to enable long chain synthesis to occur. These data provide unique insights into how heterodimeric cis-PTases have evolved from their ancestral, homodimeric forms to fulfill their function in long chain polyprenol synthesis.

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Structural elucidation of thecis-prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2008381117 ◽

2020 ◽

Vol 117 (34) ◽

pp. 20794-20802 ◽

Cited By ~ 1

Author(s):

Ban H. Edani ◽

Kariona A. Grabińska ◽

Rong Zhang ◽

Eon Joo Park ◽

Benjamin Siciliano ◽

...

Keyword(s):

Crystal Structure ◽

Active Site ◽

Protein Glycosylation ◽

Rate Limiting Step ◽

Limiting Step ◽

Chain Synthesis ◽

Rate Limiting ◽

Activity Comparison ◽

Long Chain ◽

Resolution Structure

Cis-prenyltransferase (cis-PTase) catalyzes the rate-limiting step in the synthesis of glycosyl carrier lipids required for protein glycosylation in the lumen of endoplasmic reticulum. Here, we report the crystal structure of the human NgBR/DHDDS complex, which represents an atomic resolution structure for any heterodimericcis-PTase. The crystal structure sheds light on how NgBR stabilizes DHDDS through dimerization, participates in the enzyme’s active site through its C-terminal -RXG- motif, and how phospholipids markedly stimulatecis-PTase activity. Comparison of NgBR/DHDDS with homodimericcis-PTase structures leads to a model where the elongating isoprene chain extends beyond the enzyme’s active site tunnel, and an insert within the α3 helix helps to stabilize this energetically unfavorable state to enable long-chain synthesis to occur. These data provide unique insights into how heterodimericcis-PTases have evolved from their ancestral, homodimeric forms to fulfill their function in long-chain polyprenol synthesis.

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