By the same authors

From the same journal

Activity and substrate specificity of lytic polysaccharide monooxygenases: An ATR FTIR-based sensitive assay tested on a novel species from Pseudomonas putida

Research output: Contribution to journalArticlepeer-review

Published copy (DOI)

Author(s)

  • Ilenia Serra
  • Daniele Piccinini
  • Alessandro Paradisi
  • Luisa Ciano
  • Marzia Bellei
  • Carlo Augusto Bortolotti
  • Gianantonio Battistuzzi
  • Marco Sola
  • Paul H. Walton
  • Giulia Di Rocco

Department/unit(s)

Publication details

JournalProtein science
DateAccepted/In press - 3 Dec 2021
DateE-pub ahead of print (current) - 11 Dec 2021
Number of pages11
Early online date11/12/21
Original languageEnglish

Abstract

Pseudomonas putida W619 is a soil Gram-negative bacterium commonly used in environmental studies thanks to its ability in degrading many aromatic compounds. Its genome contains several putative carbohydrate-active enzymes such as glycoside hydrolases and lytic polysaccharide monooxygenases (PMOs). In this study, we have heterologously produced in Escherichia coli and characterized a new enzyme belonging to the AA10 family, named PpAA10 (Uniprot: B1J2U9), which contains a chitin-binding type-4 module and showed activity toward β-chitin. The active form of the enzyme was produced in E. coli exploiting the addition of a cleavable N-terminal His tag which ensured the presence of the copper-coordinating His as the first residue. Electron paramagnetic resonance spectroscopy showed signal signatures similar to those observed for the copper-binding site of chitin-cleaving PMOs. The protein was used to develop a versatile, highly sensitive, cost-effective and easy-to-apply method to detect PMO's activity exploiting attenuated total reflection-Fourier transform infrared spectroscopy and able to easily discriminate between different substrates.

Bibliographical note

Funding Information:
This work was supported by the University of Modena and Reggio Emilia Finanziamento FARDSV 2019. Luisa Ciano and Paul H. Walton gratefully acknowledge the support of the UK's Biotechnology and Biological Sciences Research Council (BBSRC) through grants BB/L001926/1 and BB/L021633/1. Alessandro Paradisi and Paul H. Walton gratefully thank the Lesley Wild Scholarship for funding.

Publisher Copyright:
© 2021 The Protein Society.

    Research areas

  • activity, ATR FTIR spectroscopy, chitin, EPR, lytic polysaccharide monooxygenase

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