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Structural Organization of the Active Center of Unmodified Recombinant Sulfatase from the Mycelial Fungi Fusarium proliferatum LE1

Russian Journal of Bioorganic Chemistry volume 46, pages 563–571 (2020)Cite this article

Abstract—

Sulfatases catalyze the hydrolysis of sulfuric acid esters and play a key role in a number of biological processes of both higher eukaryotes and prokaryotes. According to literature data, for the implementation of catalysis, some representatives of this group of enzymes require posttranslational modification of serine or cysteine residues in the active center into the unique amino acid Cα-formylglycine. Nevertheless, it is confirmed that even in the absence of this modification, some sulfatases are capable of catalyzing the hydrolysis of sulfoesters. In this work, we studied the structural and functional features of active recombinant sulfatase from the mycelial fungus Fusarium proliferatum LE1, which contains a cysteine residue in the active center. A theoretical atomic model of the enzyme was first constructed, the structural organization of its active center was determined, and key amino acid residues involved in the binding of p-nitrophenyl sulfate and in the reaction of its hydrolysis were identified. Point amino acid substitutions of the identified residues led to inactivation of the enzyme. In particular, mutant forms of the enzyme with the replacement of catalytic cysteine by serine and threonine containing a hydroxyl group in the side chain completely lost activity, which indicates the direct participation of the mercapto group of cysteine in the hydrolysis reaction.

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ACKNOWLEDGMENTS

Sequencing of DNA samples was carried out using the equipment of the resource center of the Science Park of St. Petersburg State University “Development of Molecular and Cellular Technologies.”

Molecular dynamics was carried out using the computing resources of the Peter the Great Supercomputer Center of the St. Petersburg Polytechnic University (www.scc.spbstu.ru).

Funding

The study was financially supported by the Russian Foundation for Basic Research (project no. 18-34-00143).

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Authors and Affiliations

  1. Petersburg Nuclear Physics Institute, National Research Center Kurchatov Institute, 188300, Gatchina, Leningrad oblast, Russia

    N. V. Kolchina, G. N. Rychkov, A. A. Kulminskaya, F. M. Ibatullin, M. G. Petukhov & K. S. Bobrov

  2. Peter the Great St. Petersburg Polytechnic University, 195251, St. Petersburg, Russia

    N. V. Kolchina, G. N. Rychkov & A. A. Kulminskaya

  3. Granov Russian Research Center of Radiology and Surgical Technologies, 197758, Pesochnyi, St. Petersburg, Russia

    M. G. Petukhov

Authors
  1. N. V. Kolchina
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  2. G. N. Rychkov
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  3. A. A. Kulminskaya
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  4. F. M. Ibatullin
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  5. M. G. Petukhov
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  6. K. S. Bobrov
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Corresponding author

Correspondence to K. S. Bobrov.

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This work does not contain any research involving humans or animals as research objects.

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The authors declare they have no conflict of interests.

Additional information

Abbreviations: FGly, Cα-formyl glycine; F.p.Sulf-6His, Recombinant Sulfatase from Fusarium proliferatum LE1; pNPS, p-nitrophenyl sulfate; SmCS, sulfatase from Sinorhizobium meliloti; HsS, sulfatase from Homo sapiens; BtS, sulfatase from Bacteroides thetaiotaomicron; MD, molecular dynamics.

Corresponding author: e-mail: bobrov_ks@pnpi.nrcki.ru.

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Kolchina, N.V., Rychkov, G.N., Kulminskaya, A.A. et al. Structural Organization of the Active Center of Unmodified Recombinant Sulfatase from the Mycelial Fungi Fusarium proliferatum LE1. Russ J Bioorg Chem 46, 563–571 (2020). https://doi.org/10.1134/S1068162020040081

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  • DOI: https://doi.org/10.1134/S1068162020040081

Keywords:

  • sulfatase
  • Cα-formylglycine
  • posttranslational modification
  • enzyme-substrate complex