Functional metagenomics approach for discovery of novel cold-active protease from the antarctic region

Abd Latip, Muhammad Asyraf and Nordin, Noor Faizul Hadry and Alias, Siti Aisyah and Smykla, Jerzy and Yusof, Faridah and Mohamad, Mohd Azrul Naim (2024) Functional metagenomics approach for discovery of novel cold-active protease from the antarctic region. IIUM Engineering Journal, 25 (2). pp. 69-82. ISSN 1511-788X E-ISSN 2289-7860

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Abstract

The structural complexity of bacterial life makes most of it impossible to culture. Functional metagenomics approaches overcome the limitations of a culture-based approach in exploring and assessing the genetic materials of uncultured microbes. The objective of this study was to identify clones with cold-active proteases through functional metagenomics. In this work, the environmental DNA (eDNA) isolated directly from Antarctic soils was ligated into the pCC1FOS fosmid vector, transformed into EPI300-T1R E. coli host cells, and screened for proteolytic enzymes. Positive protease-producing clones were identified and isolated on skim milk agar supplemented with chloramphenicol and arabinose. This clone harbored a fosmid, pCC1FOS, which has a 48.5 kb insert that has been completely sequenced in both directions. Further analysis of the insert showed 70 NODEs. The NODE_42 encoded hypothetical protein of 297 amino acids showed a significant match to Peptidase M23 and PG-binding 1 proteins families. A three-dimensional model of the predicted protease was generated based on the known mesophilic protease of Neisseria meningitides (PDB: 3SLU). The structural alignment showed 27.07 % similarity with RMSD value of 0.402 Å based on 58 aligned residues. The active site residues were identical, but major deletions were observed in the predicted proteases. This predicted protease showed higher activity at -20 °C and 20 °C than the positive control (protease from bovine pancreas). Functional metagenomics is a promising approach in the discovery of cold-active protease with low homology to the known sequences and expressed in the host cell that has the potential for bioprospecting in low-temperature applications.

Item Type:	Article (Journal)
Additional Information:	4716/118026
Uncontrolled Keywords:	metagenomics; cold-active; enzyme; protease; Antarctic
Subjects:	Q Science > QR Microbiology T Technology > TP Chemical technology > TP248.13 Biotechnology
Kulliyyahs/Centres/Divisions/Institutes (Can select more than one option. Press CONTROL button):	International Institute for Halal Research and Training (INHART) Kulliyyah of Engineering Kulliyyah of Engineering > Department of Biotechnology Engineering Kulliyyah of Science Kulliyyah of Science > Department of Biotechnology
Depositing User:	Dr Noor Faizul Hadry Nordin
Date Deposited:	04 Jan 2025 22:43
Last Modified:	04 Jan 2025 22:43
URI:	http://irep.iium.edu.my/id/eprint/118026

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