Identification, overexpression, purification, and biochemical characterization of a novel hyperthermostable keratinase f
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ORIGINAL ARTICLE
Identification, overexpression, purification, and biochemical characterization of a novel hyperthermostable keratinase from Geoglobus acetivorans Duangjai Sittipol1 · Sudarat Rodpan1 · Ya’u S. Ajingi1,4 · Tassanee Lohnoo2 · Tassanee Lerksuthirat2 · Yothin Kumsang2 · Wanta Yingyong2 · Pongsak Khunrae1 · Triwit Rattanarojpong1 · Kovit Pattanapanyasat3 · Nujarin Jongruja1 Received: 20 August 2020 / Accepted: 30 October 2020 © King Abdulaziz City for Science and Technology 2020
Abstract The goal of this study was to identify and biochemically characterize a novel hyperthermostable keratinase from microorganisms for feather waste degradation. Here, a hyperthermophilic Geoglobus acetivorans keratinase (GacK) gene was chosen based on a search of a sequence database. The selected GacK gene was synthesized, cloned, and successfully expressed without a signal peptide in the E. coli system. A monomer of approximately 58 kDa was obtained in a soluble form and purified. The recombinant GacK displayed the highest activity at an optimum temperature of 100 °C and a pH of 10. The hyperthermostable GacK enzymatic performance remained high even after incubation in nonionic surfactants and the chelating agent EDTA. The residual and keratinolytic activities of GacK, as determined with azocasein and keratin azure used as substrates, remained significantly greater than 80% at 130 °C for 7 h. The kinetic parameters Km and Vmax for azure keratin were 0.41 mg/ml and 875.14 unit/mg, respectively, while those for azocasein were 1.51 mg/ml and 505.32 unit/mg, respectively. The results suggest that the enzyme is among the most hyperthermostable keratinases. Because of its enzymatic characteristics to degrade keratin azure at high temperatures, GacK may potentially be utilized in future industrial applications. Keywords Geoglobus acetivorans · Hyperthermostability · Industrial applications · Keratinase · Serine protease · Surfactants
* Nujarin Jongruja [email protected]
Triwit Rattanarojpong [email protected]
Duangjai Sittipol [email protected]
Kovit Pattanapanyasat [email protected]
Sudarat Rodpan [email protected]
1
Department of Microbiology, Faculty of Science, King Mongkut’s University of Technology Thonburi, 126 Pracha Uthit Rd., Bang Mod, Thung Khru, Bangkok 10140, Thailand
Tassanee Lohnoo [email protected]
2
Research Center, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok, Thailand
Tassanee Lerksuthirat [email protected]
3
Office for Research and Development, Department of Immunology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand
4
Department of Biology, Kano University of Science and Technology (KUST), Wudil, Nigeria
Ya’u S. Ajingi [email protected]
Yothin Kumsang [email protected] Wanta Yingyong [email protected] Pongsak Khunrae [email protected]
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Abbreviations GacK Geoglobus acetivor
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