Abstract
Objective
Identification and characterization of a novel thermostable amidase (Xam) with wide pH tolerance and broad-spectrum substrate specificity.
Results
Xam was identified from non-thermophilic Xinfangfangia sp. DLY26 and its acyl transfer activity was investigated. Recombinant Xam was optimally active at 60 °C and pH 9.0. The enzyme had a half life of 18 h at 55 °C and maintained more than 60 % of its maximum activity in the range of pH 3.0–11.0. Additionally, Xam exhibited broad substrate specificity towards aliphatic, aromatic, and heterocyclic amides.
Conclusions
These unique properties make Xam a promising biocatalyst for production of important hydroxamic acids at elevated temperatures.
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Supporting information
Supplementary Fig. 1—Sequence alignment of Xam with other amidases. Conserved residues are marked with black arrow. (Xam: Xinfangfangia sp. DLY26; YP_145063.1: Thermus thermophilus HB8; NP_229077.1: Thermotoga maritima MSB8; BAA36596.1: Rhodococcus sp. N-771; AET98897.1: Paracoccus sp. M-1; CAD36560.1: Rhodococcus erythropolis; WP_045444860.1: Tepidicaulis marinus).
Supplementary Fig. 2—SDS-PAGE analysis of amidase samples. M, protein marker; I, soluble extract of induced recombinant strain; II, purified Xam.
Supplementary Fig. 3—Elution profile of the purified Xam on column Superose 6 10/300 GL. 1, Ferritin (440 kDa); 2, Glucose oxidase (160 kDa); 3, BSA (66.4 kDa); 4, Albumin (45 kDa); 5, α-chymotrypsin ogen A (25.7 kDa); 6, Lysozyme (14 kDa).
Funding
This work was supported by the National Natural Science Foundation of China (Nos. 21473256, 3140005), the Key Research and Development Project of Shandong Province (Nos. 2019GSF107077, 2019GGX102062) and the Fundamental Research Funds for the Central Universities of China (No. 18CX05015A).
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Xi, L., Tan, W., Li, J. et al. Cloning and characterization of a novel thermostable amidase, Xam, from Xinfangfangia sp. DLY26. Biotechnol Lett 43, 1395–1402 (2021). https://doi.org/10.1007/s10529-021-03124-y
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DOI: https://doi.org/10.1007/s10529-021-03124-y