Purification, Characterization, and Application of Endoglucanase from Rhizopus oryzae as Antibiofilm Agent

Moumita Karmakar, Dibyajit Lahiri, Moupriya Nag, Bandita Dutta, Sudipta Dash, Tanmay Sarkar, Soumya Pandit, Vijay Jagdish Upadhye, Rina Rani Ray

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

The enzyme endoglucanase is responsible for the depolymerization of cellulose. This study focuses on characterization and purification of endoglucanase from Rhizopus oryzae MTCC 9642 through a simple size exclusion method and its effective application as an antibiofilm agent. Extracellular ß-1,4-endoglucanase, an enzyme that catalyzes the hydrolysis of carboxymethyl cellulose, was found to be synthesized by Rhizopus oryzae MTCC 9642. The enzyme was purified up to homogeneity simply by size exclusion process through ultrafiltration and gel chromatography. The molecular weight of purified enzyme protein was estimated to be 39.8 kDa and it showed the highest substrate affinity towards carboxymethyl-cellulose with Km and Vmax values of 0.833 mg ml−1 and of 0.33 mmol glucose min−1 mg−1protein, respectively. The purified enzyme exhibited optimal activity at pH 6 with a broad stability range of pH 3–8. The most preferred temperature was 35 °C and 50% of activity could be retained after the thermal exposure at 40 °C for 25 min. The purified enzyme protein was inactivated by Cu2+, while the activity could be enhanced by the addition of exogenous thiols. Since biofilm is a challenge for health sector, with the aim of eradicating the biofilm, the purified endoglucanase was used to remove biofilm produced by two nosocomial bacteria. As predicted by in silico molecular docking interaction, the purified enzyme could effectively degrade biofilm architecture of bacterial strains S. aureus and P. aeruginosa by 76.52 ± 6.52% and 61.67 ± 8.76%, respectively. The properties of purified enzyme protein, as elucidated by in vitro and in silico characterization, may be favourable for its commercial applications as a potent antibiofilm agent.

Original languageEnglish
Pages (from-to)5439-5457
Number of pages19
JournalApplied Biochemistry and Biotechnology
Volume195
Issue number9
DOIs
StatePublished - 1 Sep 2023
Externally publishedYes

Keywords

  • Antibiofilm
  • Carboxymethyl cellulose
  • Endoglucanase, Rhizopus oryzae
  • Purification

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Bioengineering
  • Molecular Biology
  • Biochemistry
  • Biotechnology

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