High-level expression and characterization of a thermostable thermolysin-like metalloprotease with industrial potential
Folia Microbiologica, 2026 (SCI-Expanded, Scopus)
- Yayın Türü: Makale / Tam Makale
- Basım Tarihi: 2026
- Doi Numarası: 10.1007/s12223-026-01564-5
- Dergi Adı: Folia Microbiologica
- Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, BIOSIS, Chemical Abstracts Core, EMBASE, MEDLINE, Natural Science Collection (ProQuest), Biological Science Database (ProQuest), Biomedical Reference Collection: Corporate Edition (EBSCO), Health Research Premium Collection (ProQuest), Pharma Collection (ProQuest)
- Anahtar Kelimeler: Characterization, Industrial enzyme applications, Purification, Recombinant metalloprotease
- Hakkari Üniversitesi Adresli: Evet
Özet
Proteases with high thermostability and catalytic efficiency are highly sought after for industrial applications, particularly within the M4 family of thermolysin-like metalloproteases. In this study, a thermostable thermolysin-like metalloprotease from Geobacillus thermoleovorans HBB208 was cloned and heterologously expressed in Escherichia coli BL21(DE3), yielding a soluble recombinant enzyme. The purified mature enzyme (GtRS1pro; ~34.6 kDa) contains the conserved HEXXH + E catalytic motif, a Zn2+-centered active site, and multiple Ca2+-binding sites characteristic of M4 proteases. GtRS1pro exhibited optimal activity at pH 8.0 and 70 °C, with a high catalytic efficiency (kcat/Km) of 2.25 × 106 M− 1 s− 1. The enzyme showed remarkable thermostability (T50 = 84.6 °C) and retained 99% residual activity after 1 h at 70 °C in the presence of 10 mM Ca2+. Functional assays demonstrated efficient hydrolysis of protein-rich substrates, including meat, collagen, and keratin. These properties position GtRS1pro as a robust and industrially relevant biocatalyst for high-temperature proteolysis and biowaste valorization, with potential applications in food processing and nutraceutical production.