Structural insight into the unique conformation of cystathionine β-synthase from Toxoplasma gondii

Fernández-Rodríguez, Carmen; Oyenarte, Iker; Conter, Carolina; González‐Recio, Irene; Núñez‐Franco, Reyes; Gil-Pitarch, Clàudia; Quintana, Iban; Jiménez‐Osés, Gonzalo; Dominici, Paola; Martínez‐Chantar, María Luz; Astegno, Alessandra; Martínez-Cruz, L.A.

doi:10.1016/j.csbj.2021.05.052

Cited by 7 publications

(24 citation statements)

References 53 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Indeed, the C-terminally truncated enzyme was eluted in SEC as one larger peak corresponding to a dimeric species and a minor peak to a tetrameric form (dimer and tetramer populations were 97% and 3%, respectively). The same behavior was previously observed for full-length TgCBS (92% dimer and 8% tetramer) ( Figure 3 A) [ 21 , 22 ]. These results were further confirmed by native PAGE analysis of TgCBS R353* at various polyacrylamide concentrations (8, 9, 10, and 12%).…”

Section: Resultssupporting

confidence: 86%

“…The same experiments were performed using L-OAS as a substrate and, again, no major differences in kinetic parameters between the full-length and truncated enzyme were found ( Table 1 ). Moreover, the addition of L-Ser or L-OAS resulted in the disappearance of the internal aldimine band (411 nm) and the concomitant appearance of the aminoacrylate band (460 nm) in both absorbance and CD spectra ( Figure 4 C,D), as already observed for the full-length TgCBS [ 21 , 22 ].…”

Section: Resultssupporting

confidence: 75%

“…The ΔH of the peak at 39 °C closely matches the theoretical ΔH value for linker region unfolding (19.5 kcal/mol). A close view of the crystal structure of TgCBS [ 22 ] shows that the interdomain linker folds on itself and is fixed by a combined set of interactions at its two extremes, with both the catalytic domain and the Bateman module ( Figure 6 D). Beyond reducing the mobility of the linker, these contacts help to configure a sole protein structural body with almost no mobile elements, which is consistent with a single peak in the thermal denaturation event in the full-length TgCBS protein.…”

Section: Resultsmentioning

confidence: 99%

“…Beyond reducing the mobility of the linker, these contacts help to configure a sole protein structural body with almost no mobile elements, which is consistent with a single peak in the thermal denaturation event in the full-length TgCBS protein. Interestingly, and despite sharing a similar overall basket-like fold in the absence of AdoMet, the HsCBS [ 22 ] shows a less restricted linker, an essential feature to allow the displacement of the Bateman module when binding the allosteric activator.…”

Section: Resultsmentioning

confidence: 99%

“…In contrast to HsCBS, TgCBS does not possess an N-terminal heme binding region and is unresponsive to AdoMet, despite containing a C-terminal Bateman module ( Figure 1 ). Notably, the three-dimensional structure of a TgCBS construct missing a few residues from an internal loop (which are not involved in the activity or the oligomerization state, TgCBS Δ466–491) revealed that the enzyme exists in an active basal-type (basket-shaped) folding, which likely constitutes its sole conformational state [ 22 ]. These findings raise critical new questions about whether CBSs with similar assemblies are always (or not) subject to evolve allosterically towards a second activated conformation.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Insights into Domain Organization and Regulatory Mechanism of Cystathionine Beta-Synthase from Toxoplasma gondii

Conter

Fruncillo

Favretto

et al. 2022

IJMS

Self Cite

View full text Add to dashboard Cite

Cystathionine beta-synthase (CBS) is a key regulator of homocysteine metabolism. Although eukaryotic CBS have a similar domain architecture with a catalytic core and a C-terminal Bateman module, their regulation varies widely across phyla. In human CBS (HsCBS), the C-terminus has an autoinhibitory effect by acting as a cap that avoids the entry of substrates into the catalytic site. The binding of the allosteric modulator AdoMet to this region alleviates this cap, allowing the protein to progress from a basal toward an activated state. The same activation is obtained by artificial removal or heat-denaturation of the Bateman module. Recently, we reported the crystal structure of CBS from Toxoplasma gondii (TgCBS) showing that the enzyme assembles into basket-like dimers similar to the basal conformers of HsCBS. These findings would suggest a similar lid function for the Bateman module which, as in HsCBS, should relax in the absence of the C-terminal module. However, herein we demonstrate that, in contrast with HsCBS, removal of the Bateman module in TgCBS through deletion mutagenesis, limited proteolysis, or thermal denaturation has no effects on its activity, oligomerization, and thermal stability. This opposite behavior we have now found in TgCBS provides evidence of a novel type of CBS regulation.

show abstract

Section: Resultssupporting

confidence: 86%

Section: Resultssupporting

confidence: 75%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Insights into Domain Organization and Regulatory Mechanism of Cystathionine Beta-Synthase from Toxoplasma gondii

Conter

Fruncillo

Favretto

et al. 2022

IJMS

Self Cite

View full text Add to dashboard Cite

show abstract

Design, synthesis, and bioactivity evaluation of clindamycin derivatives against multidrug‐resistant bacterial strains

Jia,

Zhang,

Zhu

et al. 2024

Drug Development Research

View full text Add to dashboard Cite

Our research aims to reduce the bacterial resistance of clindamycin against Gram‐positive bacteria and expand its range of bacterial susceptibility. First, we optimized the structure of clindamycin based on its structure–activity relationship. Second, we employed the fractional inhibitory concentration method to detect drugs suitable for combination with clindamycin derivatives. We then used a linker to connect the clindamycin derivatives with the identified combined therapy drugs. Finally, we tested antibacterial susceptibility testing and conducted in vitro bacterial inhibition activity assays to determine the compounds. with the highest efficacy. The results of our study show that we synthesized clindamycin propionate derivatives and clindamycin homo/heterodimer derivatives, which exhibited superior antibacterial activity compared to clindamycin and other antibiotics against both bacteria and fungi. In vitro bacteriostatic activity testing against four types of Gram‐negative bacteria and one type of fungi revealed that all synthesized compounds had bacteriostatic effects at least 1000 times better than clindamycin and sulfonamides. The minimum inhibitory concentration (MIC) values for these compounds ranged from 0.25 to 0.0325 mM. Significantly, compound 5a demonstrated the most potent inhibitory activity against three distinct bacterial strains, displaying MIC values spanning from 0.0625 to 0.0325 mM. Furthermore, our calculations indicate that compound 5a is safe for cellular use. In conclusion, the synthesized compounds hold great promise in addressing bacterial antibiotic resistance.

show abstract

Structural basis of the inhibition of cystathionine γ‐lyase from Toxoplasma gondii by propargylglycine and cysteine

et al. 2023

Self Cite

View full text Add to dashboard Cite

Cystathionine γ‐lyase (CGL) is a PLP‐dependent enzyme that catalyzes the last step of the reverse transsulfuration route for endogenous cysteine biosynthesis. The canonical CGL‐catalyzed process consists of an α,γ‐elimination reaction that breaks down cystathionine into cysteine, α‐ketobutyrate, and ammonia. In some species, the enzyme can alternatively use cysteine as a substrate, resulting in the production of hydrogen sulfide (H2S). Importantly, inhibition of the enzyme and consequently of its H2S production activity, makes multiresistant bacteria considerably more susceptible to antibiotics. Other organisms, such as Toxoplasma gondii, the causative agent of toxoplasmosis, encode a CGL enzyme (TgCGL) that almost exclusively catalyzes the canonical process, with only minor reactivity to cysteine. Interestingly, the substitution of N360 by a serine (the equivalent amino acid residue in the human enzyme) at the active site changes the specificity of TgCGL for the catalysis of cystathionine, resulting in an enzyme that can cleave both the CγS and the CβS bond of cystathionine. Based on these findings and to deepen the molecular basis underlying the enzyme‐substrate specificity, we have elucidated the crystal structures of native TgCGL and the variant TgCGL‐N360S from crystals grown in the presence of cystathionine, cysteine, and the inhibitor d,l‐propargylglycine (PPG). Our structures reveal the binding mode of each molecule within the catalytic cavity and help explain the inhibitory behavior of cysteine and PPG. A specific inhibitory mechanism of TgCGL by PPG is proposed.

show abstract

Structural insight into the unique conformation of cystathionine β-synthase from Toxoplasma gondii

Abstract: Graphical abstract

Cited by 7 publications

References 53 publications

Insights into Domain Organization and Regulatory Mechanism of Cystathionine Beta-Synthase from Toxoplasma gondii

Insights into Domain Organization and Regulatory Mechanism of Cystathionine Beta-Synthase from Toxoplasma gondii

Design, synthesis, and bioactivity evaluation of clindamycin derivatives against multidrug‐resistant bacterial strains

Structural basis of the inhibition of cystathionine γ‐lyase from Toxoplasma gondii by propargylglycine and cysteine

Contact Info

Product

Resources

About