Regular Article
Identification of a Carboxylesterase as the Major Protein Bound by Molinate,☆☆

https://doi.org/10.1006/taap.1998.8381Get rights and content

Abstract

Molinate, a herbicide widely used on rice, has been previously shown to cause testicular toxicity when a single dose is administered to Sprague–Dawley rats. The sulfoxide metabolite of molinate also was capable of eliciting testicular damage but at lower dose levels than molinate, suggesting that metabolic activation via sulfoxidation could be important in testicular toxicity. Both the sulfoxide and sulfone metabolites of molinate are electrophilic and molinate covalent binding to cellular macromolecules has been attributed to formation of these reactive metabolites. The present study has investigated the nature of the binding reaction of14C-molinate as well as14C-molinate sulfoxide and14C-molinate sulfone in liver and testis microsomal preparations. All three compounds in preparations from both tissues bound extensively and tightly to only one protein of approximately 60 kDa molecular weight on SDS–PAGE. Isoelectric focusing PAGE revealed a pI of approximately 6.0 and native PAGE analysis revealed a native molecular weight of 180 kDa. These data, along with the ability of phenylmethylsulfonyl fluoride to block binding of the14C-molinate, suggested the molinate-bound protein was an esterase. The protein was purified to homogeneity and MALDI-TOF mass spectral analysis was consistent with Hydrolase A, a carboxylesterase present in both liver and testis. N-terminal sequence analysis revealed 100% homology with Hydrolase A for the first 17 residues. The effect of molinate administration onin vivoesterase activity was assessed both by enzymatic measurement and by histochemical measurement. Molinate treatment caused a marked inhibition of nonspecific esterase activity in both liver and testis. In the testis, histochemical staining showed the esterase activity inhibited by molinate was localized primarily to the Leydig cell, consistent with the localization of Hydrolase A. From these data, it is proposed that molinate-induced inhibition of esterase activity in the Leydig cell could inhibit the mobilization of cholesterol esters required for testosterone biosynthesis.

References (32)

  • B. Yan et al.

    Rat testicular carboxylesterase: Cloning, cellular localization and relationship to liver Hydrolase A

    Arch. Biochem. Biophys.

    (1995)
  • R. Arndt et al.

    Purification and molecular properties of an unspecific carboxylesterase from rat-liver microsomes

    Eur. J. Biochem.

    (1973)
  • A.W. Blackshaw

    Localization of testicular enzymes

    The Testis, II: Biochemistry

    (1970)
  • Cal-EPA, 1996, Molinate (Ordram): Risk Characterization Document, Department of Pesticide Regulation,...
  • J.R. DeBaun et al.

    Metabolism of14

    J. Agric. Food Chem.

    (1978)
  • Cited by (47)

    • Label-free, rapid, and sensitive detection of carboxylesterase using surfactant-doped liquid crystal sensor

      2019, Journal of Molecular Liquids
      Citation Excerpt :

      As reported, nephritis is caused by protein-overload proteinuria [36]. As a CES-like urinary excreted protein [37], the detection of CES in human urine could assist the diagnosis of nephritis. It can be seen from Fig. S8, a higher concentration of urine (e.g. 100%, 50%) could affect the LC-based sensing substrate.

    • The Leydig Cell as a Target for Toxicants

      2018, Comprehensive Toxicology: Third Edition
    • Russian VX

      2015, Handbook of Toxicology of Chemical Warfare Agents: Second Edition
    • Effects of selected xenobiotics on hepatic and plasmatic biomarkers in juveniles of Solea senegalensis

      2014, Environmental Research
      Citation Excerpt :

      This result contrasts with no effect over hepatic CbE (pNPA) in male guppies after water exposures up to 300 µg/l (Li, 2008). The physiological role of CbEs is still under debate but they are related to lipid metabolism and steroidogenesis (Jewell and Miller, 1998), thus any action over their activity would likely have significant physiological consequences. In addition, CbE and MO activities responded similarly as they both coincided on the action over phase I enzymes after injections with βNF (induction), keto (inhibition) and of mixed nature after xenoestrogens injection (induction and inhibition depending on the form).

    • Acetylcholinesterase activity in the terrestrial snail Xeropicta derbentina transplanted in apple orchards with different pesticide management strategies

      2011, Environmental Pollution
      Citation Excerpt :

      Attempts to establish a relationship between inhibition of AChE activity and toxic effects at higher levels of biological organization has been a matter of growing concern in ecotoxicology. Histological damage (Jewell and Miller, 1998; Espinoza-Navarro and Bustos-Obregón, 2005), physiological and metabolic changes (Grue et al., 1997) and behavioural perturbations (Burger et al., 2002; Scott and Sloman, 2004) have been the major toxicological endpoints associated to inhibition of nervous AChE activity in aquatic and vertebrate organisms. However, comparable investigations with terrestrial snails are still limited.

    • The Leydig Cell as a Target for Toxicants

      2010, Comprehensive Toxicology, Second Edition
    View all citing articles on Scopus

    Supported by California Rice Research Board (RP-7) and by National Institute of Environmental Health Sciences Training Grant 5-T32-EOS-7059.

    ☆☆

    Johnson, A. D.

    View full text