Xenopus laevis and human type 3 iodothyronine deiodinase enzyme cross-species sensitivity to inhibition by ToxCast chemicals

Mayasich, Sally A.; Korte, Joseph J.; Denny, Jeffrey S.; Hartig, Phillip C.; Olker, Jennifer H.; DeGoey, Philip; O’Flanagan, Joseph; Degitz, Sigmund J.; Hornung, Michael W.

doi:10.1016/j.tiv.2021.105141

Cited by 11 publications

(13 citation statements)

References 50 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Hill slopes for the concentration-response curves in the present study did not differ significantly among variants for most chemicals (Figure 3), but steeper slopes for some chemicals are a potential indication of binding to more than one site (allosteric inhibition; Prinz, 2010;Shoichet, 2006). The steeper Hill slopes are clues to multisite binding that had also been observed in previous cross-species screening assays, specifically for ascorbyl palmitate, bisphenol A diglycidyl ether, and kepone (Mayasich et al, 2021). Therefore, the MOE software Site Finder tool was employed to explore potential sites for cofactor binding.…”

Section: Discussionsupporting

confidence: 71%

“…Notably, all of the nonhalogenated chemicals tested in the present study (ascorbyl palmitate, bisphenol A diglycidyl ether, nordihydroguaiaretic acid, and XTH) are predicted to prefer the cofactor site for the A240R and C239S variant models compared with a catalytic site preference for the other variant models. Kepone and all of these chemicals except XTH showed significant differences in concentration–response curves between hDIO3 and Xldio3 in a previous study (Mayasich et al, 2021).…”

Section: Resultsmentioning

confidence: 55%

“…The change from alanine to arginine in variant A240R differed from WT for only one chemical (Figure 2 and Table 6), indicating that arginine at Position 240 has little or no influence on perturbation of deiodinase activity by the chemicals tested in the present study. Therefore, we can likely reject the hypothesis that arginine at 240 was a critical amino acid responsible for significant differences between Xldio3 and hDIO3 concentration-response results in a previous study (Mayasich et al, 2021).…”

Section: Discussionmentioning

confidence: 65%

“…From previous substrate preference comparisons (Mayasich et al, 2021), the highest activity of hDIO3 was observed toward T3. Therefore, the mean T3 activity as net change in A420 (net dA420) was taken first for each variant; then, the expressed T4 activity measurements were calculated as percentage of the mean T3 activity for the relevant variant.…”

Section: Methodsmentioning

confidence: 68%

“…In the previous assays for the Xldio3 enzyme, the model inhibitor XTH had a mean IC50 across all 12 screening-plate runs of 0.31 ± 0.06 µM, with a range of 0.26 to 0.42 µM, Hill slope mean −1.01 ± 0.12, range −0.81 to −1.19 (see Supplementary Table 5 in Mayasich et al, 2021). Olker et al (2019) reported a mean IC50 for hDIO3 across all 48 runs of 0.3 ± 0.1 µM, with a range of 0.14 to 0.46 µM, and Hill slope mean −0.9 ± 0.1, range −0.72 to −1.08.…”

Section: In Vitro Deiodinase Inhibitionmentioning

confidence: 98%

See 4 more Smart Citations

Combining In Vitro and In Silico New Approach Methods to Investigate Type 3 Iodothyronine Deiodinase Chemical Inhibition Across Species

Mayasich

Goldsmith

Mattingly

et al. 2023

Enviro Toxic and Chemistry

Self Cite

View full text Add to dashboard Cite

New approach methodologies (NAMs) are being developed to reduce and replace vertebrate animal testing in support of ecotoxicology and risk assessment. The US Environmental Protection Agency's Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) bioinformatic tool was used to evaluate amino acid sequence conservation of the type 3 iodothyronine deiodinase (DIO3) enzyme across species to demonstrate NAM applications for understanding effects of chemical interactions with a specific protein target. Existing literature was used to identify critical amino acids for thyroid hormone binding and interaction with a reducing cofactor. The SeqAPASS tool identifies whether known critical amino acids involved in ligand binding are exact, partial, or not matches across species compared with a template species based on molecular weight and side chain classification. This evaluation guided the design of variant proteins representing critical amino acid substitutions found in various species. Site‐directed mutagenesis of the wild‐type (WT) human DIO3 gene sequence was used to create six variant proteins expressed in cell culture, which were then tested in vitro for chemical inhibition. Significant differences in in vitro median inhibitory concentration results were observed among variants for potential competitive inhibitors. A molecular model representing the WT human DIO3 was constructed using Molecular Operating Environment (MOE) software and mutated in silico to create the six variants. The MOE Site Finder tool identified the proposed catalytic and cofactor sites and potential alternative binding sites. Virtual docking did not provide affinity scores with sufficient resolution to rank the potency of the chemical inhibitors. Chemical characteristics, function and location of substituted amino acids, and complexities of the protein target are important considerations in developing NAMs to evaluate chemical susceptibility across species. Environ Toxicol Chem 2023;42:1032–1048. © 2023 University of Wisconsin‐Madison. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

show abstract