Inactivation of GDP-mannose dehydrogenase from Pseudomonas aeruginosa by penicillic acid identifies a critical active site loop

Kimmel, Jennifer; Tipton, Peter A.

doi:10.1016/j.abb.2005.06.028

Cited by 8 publications

(5 citation statements)

References 11 publications

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“…Pivotal to the alginate biosynthetic pathway is GDP- d -mannose dehydrogenase (GMD), which is required to form the alginate sugar nucleotide feedstock, guanosine diphosphate mannuronic acid (GDP- d -ManA). − Since there is no corresponding enzyme in humans, specific inhibition of GMD could be envisaged as a tactic to stop alginate production in chronic, mucoid P. aeruginosa infections . GMD belongs to a small group of NAD + -dependent four-electron-transfer dehydrogenases , and converts GDP- d -Man 1 to GDP- d -ManA 2 (Figure a); the NAD + -mediated oxidation is proposed to have four discrete steps, facilitated by an active site Cys268 residue .…”

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confidence: 99%

“…5−7 Since there is no corresponding enzyme in humans, specific inhibition of GMD could be envisaged as a tactic to stop alginate production in chronic, mucoid P. aeruginosa infections. 8 GMD belongs to a small group of NAD + -dependent four-electron-transfer dehydrogenases 9,10 and converts GDP-D-Man 1 to GDP-D-ManA 2 (Figure 1a); the NAD + -mediated oxidation is proposed to have four discrete steps, facilitated by an active site Cys268 residue. 11 We recently reported the first series of C6-modified sugar nucleotides to investigate this oxidative mechanism, establishing evidence of a C6-ketone intermediate from oxidation of a C6-CH 3 modified analogue.…”

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Inhibition of the GDP-d-Mannose Dehydrogenase from Pseudomonas aeruginosa Using Targeted Sugar Nucleotide Probes

et al. 2020

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Alginate, an anionic polysaccharide composed of acetylated, β-1,4 linked Dmannuronic acid and α-1,4 linked L-guluronic acid residues, constitutes the major exopolysaccharide component of mucoid Pseudomonas aeruginosa infections. Such infections are particularly deleterious for cystic fibrosis patients, 1 contributing to the establishment of a biofilm environment and augmenting the antibiotic resistance profile of the bacterium. The molecular genetics and regulation of alginate biosynthesis in P. aeruginosa have been wellcharacterised. 2,3 However, our understanding of the chemical biology underpinning polysaccharide production and modification is still incomplete. This represents a significant opportunity to comprehend and exploit such pathways, especially within the global obligation to develop new anti-bacterial strategies; epitomised by recent work developing small molecule inhibitors for c-di-GMP binding to the receptor protein Alg44, which activates alginate production in P. aeruginosa. 4 Pivotal to the alginate biosynthetic pathway is GDP-D-mannose dehydrogenase (GMD), which is required to form the alginate sugar nucleotide feedstock, guanosine diphosphate mannuronic acid (GDP-D-ManA). 5,6 Since there is no corresponding enzyme in humans, specific inhibition of GMD could be envisaged as a tactic to stop alginate production in chronic, mucoid P. aeruginosa infections. 7 GMD belongs to a small group of NAD + -dependent fourelectron-transfer dehydrogenases, 8,9 and converts GDP-D-Man 1 to GDP-D-ManA 2 (Figure 1a); the NAD + -mediated oxidation is proposed to have four discrete steps, facilitated by an active site Cys 268 residue. 10 We recently reported the first series of C6-modified sugar nucleotides to investigate this oxidative mechanism, establishing evidence of a C6-ketone intermediate from oxidation of a C6-CH3 modified analogue. 11 Pursuant of a substrate-based inhibitor of GMD and to further expand structure-activity relationship knowledge (to underpin future small molecule inhibition strategies), we report herein the design, synthesis and evaluation of a matrix of C4-, C5-and C6-modified GDP-D-Man analogues (Figure 1b). Our strategy focused on analogues of the pyranose component within the sugar nucleotide, specifically: i) C6-OH replacements, removing capability for oxidation, but retaining the potential to bind GMD ii) C6 modifications to mimic the product carboxylic acid in 2 and iii) C4 and C5 functional group changes to probe steric and electronic effects involved in substrate binding.

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Inhibition of the GDP-d-Mannose Dehydrogenase from Pseudomonas aeruginosa Using Targeted Sugar Nucleotide Probes

et al. 2020

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“…1b ), which contains a conjugated Michael acceptor, was an irreversible, nonspecific inactivator of GMD in vitro , with low selectivity for the active site of the protein. 18 In addition, the movement of the GDP-mannose binding site loop was demonstrated to be partially rate-limiting, with the interaction of Cys213-Asn252 holding the loop in a closed position during catalysis. Mutation of Cys213 to Ala213 to allow free movement of the loop was demonstrated to result in a 1.8-fold increase in V max .…”

Section: Introductionmentioning

confidence: 99%

Virtual screening, identification and in vitro validation of small molecule GDP-mannose dehydrogenase inhibitors

Dolan,

Ahmadipour,

Wahart

et al. 2023

RSC Chem. Biol.

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“…S1A). In addition, the Bcer98_2076 also shares 38% sequence identity with functional GDP‐mannose dehydrogenase from Pseudomonas aeruginosa [17]. Similarly, the potential epimerase from B. cereus , Bcer98_2077, shares 36% and 32% sequence identity with functional UGlcAE (AtUGlcAE3) from plants, and LpsL from Sinorizobium (SMc02640), respectively (Fig.…”

Section: Resultsmentioning

confidence: 99%

Biosynthesis of UDP‐glucuronic acid and UDP‐galacturonic acid in Bacillus cereus subsp. cytotoxis NVH 391‐98

Broach¹,

Gu²,

Bar‐Peled³

2011

The FEBS Journal

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SUMMARY The food borne pathogen, Bacillus cereus, produces uronic acid-containing glycans that are secreted in a shielding biofilm environment, and certain alkaliphilic Bacillus deposit uronate-glycan polymers in the cell wall when adapting to alkaline environments. The source of these acidic sugars is unknown, and here we have described the functional identification of an operon in B. cerues subsp. cytotoxis NVH 391–98 that comprises genes involved in the synthesis of UDP-uronic acids in Bacillus spp. Within the operon, a UDP-glucose 6-dehydrognease (UGlcDH) converts UDP-glucose in the presence of NAD+ to UDP-glucuronic acid and NADH, and a UDP-GlcA 4-epimerase (UGlcAE) converts UDP-glucuronic acid to UDP-galacturonic acid. Interestingly, in vitro both enzymes can utilize the TDP-sugar forms as well, albeit at lower catalytic efficiency. Unlike most of the very few bacterial 4-epimerases that have been characterized, which are promiscuous, the B. cereus UGlcAE enzyme is very specific and cannot use UDP-Glc, UDP-GlcNAc, UDP-GlcNAcA or UDP-Xyl as substrates. Size exclusion chromatography suggests that UGlcAE is active as a monomer, unlike the dimeric form of plant enzymes; the Bacillus UGlcDH is also found as a monomer. Phylogenic analysis further suggests that the Bacillus UGlcAE may have evolved separately from other bacterial and plant epimerases. Our results provide insight into the formation and function of uronic acid-containing glycans in the lifecycle of B. cereus and related species containing homologous operons as well as the basis to determine the importance of these acidic glycans. We also discuss the ability to target UGlcAE as a drug candidate.

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Inactivation of GDP-mannose dehydrogenase from Pseudomonas aeruginosa by penicillic acid identifies a critical active site loop

Cited by 8 publications

References 11 publications

Inhibition of the GDP-d-Mannose Dehydrogenase from Pseudomonas aeruginosa Using Targeted Sugar Nucleotide Probes

Inhibition of the GDP-d-Mannose Dehydrogenase from Pseudomonas aeruginosa Using Targeted Sugar Nucleotide Probes

Virtual screening, identification and in vitro validation of small molecule GDP-mannose dehydrogenase inhibitors

Biosynthesis of UDP‐glucuronic acid and UDP‐galacturonic acid in Bacillus cereus subsp. cytotoxis NVH 391‐98

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