Allosteric inhibition and kinetic characterization of Klebsiella pneumoniae CysE: An emerging drug target

Verma, Deepali; Gupta, Sunita; Saxena, Rajiv; Kaur, Punit; Rachana, R; Srivastava, Sudha; Gupta, Vibha

doi:10.1016/j.ijbiomac.2019.10.170

Cited by 10 publications

(10 citation statements)

References 47 publications

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“…L-cysteine competes directly with L-serine for the active site displaying competitive inhibition [30]. Analysis of co-crystallized structures of SAT with L-cysteine, shows L-cysteine bound in the L-serine binding pocket located in the SAT active site [6,[31][32][33], supporting the observation of L-cysteine being a competitive inhibitor relative to L-serine [5,30]. This mechanism of inhibition appears to be well-conserved across the SAT family [30,34].…”

Section: Introductionmentioning

confidence: 85%

Serine acetyltransferase from Neisseria gonorrhoeae; structural and biochemical basis of inhibition

Oldham

Prentice

Summers

et al. 2022

Biochemical Journal

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Serine acetyltransferase (SAT) catalyzes the first step in the two-step pathway to synthesize L-cysteine in bacteria and plants. SAT synthesizes O-acetylserine from substrates L‑serine and acetyl coenzyme A and is a key enzyme for regulating cellular cysteine levels by feedback inhibition of L-cysteine, and its involvement in the cysteine synthase complex. We have performed extensive structural and kinetic characterization of the SAT enzyme from the antibiotic-resistant pathogen Neisseria gonorrhoeae. Using X-ray crystallography, we have solved the structures of NgSAT with the non-natural ligand, L-malate (present in the crystallization screen) to 2.01 Å and with the natural substrate L-serine (2.80 Å) bound. Both structures are hexamers, with each monomer displaying the characteristic left-handed parallel β-helix domain of the acyltransferase superfamily of enzymes. Each structure displays both extended and closed conformations of the C-terminal tail.  L‑malate bound in the active site results in an interesting mix of open and closed active site conformations, exhibiting a structural change mimicking the conformation of cysteine (inhibitor) bound structures from other organisms. Kinetic characterization shows competitive inhibition of L-cysteine with substrates L-serine and acetyl coenzyme A. The SAT reaction represents a key point for the regulation of cysteine biosynthesis and controlling cellular sulfur due to feedback inhibition by L-cysteine and formation of the cysteine synthase complex. Data presented here provide the structural and mechanistic basis for inhibitor design and given this enzyme is not present in humans could be explored to combat the rise of extensively antimicrobial-resistant N. gonorrhoeae.

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Section: Introductionmentioning

confidence: 85%

Serine acetyltransferase from Neisseria gonorrhoeae; structural and biochemical basis of inhibition

Oldham

Prentice

Summers

et al. 2022

Biochemical Journal

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“…Members of this family are defined by a six-peptide tandem repeat, [LIV]-[GAED]-X 2 -[STAV]-X, which gives rise to a distinctive left-handed beta helix (LβH) [ 36 ]. Structural characterization of CysE enzymes with and without substrates and cysteine (inhibitor) bound from a range of Gram-negative bacterial pathogens including E. coli (1T3D) [ 37 ], H. influenzae (1SSM, 1SSQ, 1SST) [ 38 ], Yersinia pestis (3GVD) [ 39 ], Brucella melitensis (3MC4), Vibrio cholerae (4H7O), Brucella abortus (4HZC, 4HZD) [ 40 ], K. pneumoniae (6JVU) [ 41 ], N. gonorrhoeae (6WYE, 7RA4) [ 42 ] and S. typhimurium (7E3Y). These structures provide insight into active site architecture upon substrate and inhibitor binding, which can be used to inform inhibitor design.…”

Section: Structural Characteristics Of Cysementioning

confidence: 99%

“…Several promising CysE inhibitors have been identified via in silico screening of natural compound libraries. Recently, the flavonoid quercetin was found to inhibit CysE from K. pneumoniae (IC 50 = 3.7 μM) ( Table 1 ) [ 41 ]. Through docking analysis, quercetin ( Figure 3 ) was shown to bind allosterically to the CysE trimer–trimer interface.…”

Section: Natural Compound Inhibitorsmentioning

confidence: 99%

“…Through docking analysis, quercetin ( Figure 3 ) was shown to bind allosterically to the CysE trimer–trimer interface. Although not experimentally investigated by the authors, the binding of quercetin to this interface may inhibit Kp CysE through disrupting the trimer–trimer interactions, dissociating the hexamer, which has been shown to reduce CysE activity [ 41 ]. However, quercetin has been shown to inhibit other bacterial enzymes including isocitrate lyase [ 51 ] and glutamate racemase [ 52 ].…”

Section: Natural Compound Inhibitorsmentioning

confidence: 99%

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Combatting antimicrobial resistance via the cysteine biosynthesis pathway in bacterial pathogens

et al. 2022

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Antibiotics are the cornerstone of modern medicine and agriculture, and rising antibiotic resistance is one the biggest threats to global health and food security. Identifying new and different druggable targets for the development of new antibiotics, is absolutely crucial to overcome resistance. Adjuvant strategies that either enhance the activity of existing antibiotics or improve clearance by the host immune system provide another mechanism to combat antibiotic resistance. Targeting a combination of essential and non-essential enzymes that play key roles in bacterial metabolism, is a promising strategy to develop new antimicrobials and adjuvants, respectively. The enzymatic synthesis of L-cysteine is one such strategy. Cysteine plays a key role in proteins and is crucial for the synthesis of many biomolecules important for defense against the host immune system. Cysteine synthesis is a two-step process, catalyzed by two enzymes. Serine acetyltransferase (CysE) catalyzes the first step to synthesize the pathway intermediate O-acetylserine, and O-acetylserine sulfhydrylase (CysK/CysM) catalyzes the second step using sulfide or thiosulfate to produce cysteine. Disruption of the cysteine biosynthesis pathway results in dysregulated sulfur metabolism, altering the redox state of the cell leading to decreased fitness, enhanced susceptibility to oxidative stress and increased sensitivity to antibiotics. In this review we summarize the structure and mechanism of characterized CysE and CysK/CysM enzymes from a variety of bacterial pathogens, and the evidence that supports targeting these enzymes for the development of new antimicrobials or antibiotic adjuvants. In addition, we explore and compare compounds identified thus far that target these enzymes.

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“…The 3-dimensional crystal structure (PDB-ID: 2Q3C) (Schnell et al, 2007) along with some inhibitors have been reported for M.tb CysK enzyme (Poyraz et al, 2013;Ullas et al, 2013), but other than a model structure (Gupta and Gupta, 2020), mycobacterial CysE has not been characterized structurally and functionally till date. Nevertheless, the crystal structures of CysEs from other homologues such as Escherichia coli (PDB-ID:1T3D) (Pye et al, 2004), Haemophilus influenzae (PDB-ID:1S80,1SSQ) (Gorman and Shapiro, 2004;Olsen et el., 2004), Entamoeba histolytica (PDB-ID:3P1B) (Kumar et al, 2011), Brucella abortus (PDB-ID:4HZC) (Kumar et al, 2014), Brucella melitensis (PDB-ID:3MC4), Vibrio cholerae (PDB-ID:4H7O), Yersinia pestis (PDB-ID:3GVD), Bacteroides vulgates (PDB-ID:3F1X) Glycine max (PDB-ID:4N69) (Yi et al, 2013) and Klebsiella pneumonia (PDB-ID:6JVU) (Verma et al, 2020) contribute to structure function understanding of this gateway enzyme in the de novo cysteine biosynthetic pathway. CysEs exist as hexamer except for E. histolytica which supports a trimeric form (Kumar et al, 2011), and B.abortus which exists in both hexameric and trimeric forms (Kumar et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

Unveiling the structural features of CysE: a novel target for therapeutic interventions against persistent mycobacteria

Gupta

2022

JANS

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World Health Organization (WHO) reports that one-third of the world’s population is infected with a persistent form of Mycobacterium tuberculosis (M.tb), the causative bacterium responsible for causing the dreaded tuberculosis disease. Targeting mycobacterial persisters is important for achieving WHO’s End TB target. The de-novo cysteine biosynthetic pathway is a novel target for addressing M.tb persistence. The two-step pathway comprises of serine acetyltransferase/CysE and O-acetyl-serine-sulfhydrylase/OASS/CysK. The present study is an attempt to understand the structural features of mycobacterial CysE by investigating the divergence amongst orthologous through phylogenetic analysis. Mapping of mycobacterial CysE sequences on the whole orthologous (COG1045) tree segregated the species into four clusters and several isoforms leading to their descendants identification. Interestingly the analysis revealed that the extended C-terminal α-helix believed unique to M.tb is also present in other organisms such as: Campylobacter ureolyticus, Bacillus cereus, Geminocystis herdmanii and Paenibacillus borealis. Further, the Hidden Markov model search against the whole Uniprot database suggests a plausible role of C-terminal α-helix of CysE in strengthening the substrate and/or co-factor binding. In addition, phylogenetic analysis of CysE sequences from the Mycobacteriaceae family facilitates grouping them under ten well-formed and six monophyletic clades, each based on characteristic features with respect to domain architecture, oligomeric assembly, C-terminal tetra-peptide tail, regulatory and feedback mechanism etc. Employing molecular phylogeny in conjunction with structural analysis has provided detailed insights for mycobacterial CysEs as drug target.

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Allosteric inhibition and kinetic characterization of Klebsiella pneumoniae CysE: An emerging drug target

Cited by 10 publications

References 47 publications

Serine acetyltransferase from Neisseria gonorrhoeae; structural and biochemical basis of inhibition

Serine acetyltransferase from Neisseria gonorrhoeae; structural and biochemical basis of inhibition

Combatting antimicrobial resistance via the cysteine biosynthesis pathway in bacterial pathogens

Unveiling the structural features of CysE: a novel target for therapeutic interventions against persistent mycobacteria

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