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THERMUS THERMOPHILUS THERMOPHILIC ESTERASE
WO09156532
The invention relates to a thermophilic protein with esterase activity. Said protein, of the Thermus thermophilus organism, is expressed in recombinant yeasts and bacteria, in particular E.coli, K.lactis and S.cerevisiae. The esterase is expressed in fusion protein form with a signal sequence that enables secretion to the culture medium and, in addition, is operably linked to an inducible promoter. The recombinant strains are advantageous in that: (i) the cultivation thereof is much easier than that of the thermophilic organism that produces the enzyme, and (ii) they can be used to obtain a larger number of enzyme units per litre of culture than the original thermophilic strains.
Advantages / Differential Points Thermophilic esterase from Thermus thermophilus: polypeptide with esterase activity; variants and fusion constructs are covered. Thermostability at high temperatures: the esterase is stated to be thermophilic and stable between 60–90 °C (claim 3). Recombinant production in mesophiles: expression demonstrated/claimed in bacteria and yeasts—specifically E. coli, K. lactis, S. cerevisiae—with inducible promoters. Secretion to culture medium via signal peptides (e.g., α‑mating factor signals), facilitating downstream recovery. Affinity tags for purification (e.g., Flag‑tag, His‑tag) are contemplated to streamline isolation. Process benefit explicitly stated: recombinant strains are easier to cultivate than the source thermophile and allow more enzyme units per liter than original thermophilic strains. Construct variants for secretion: N‑terminal truncations (removal of 16 or 26 aa) improve secreted activity in yeasts; reported activities include ~500 U/L (longer variant) and ~200 U/L (shorter variant) in K. lactis at ~49 h, and ~1000 U/L in S. cerevisiae at ~72 h; purification factor ~30.2 after affinity chromatography. Other Applications Detergent compositions (use of the esterase in detergent formulations). Modification of fats or oils (contacting the enzyme with a fat/oil under hydrolytic conditions). Obtaining enantiopure products (use of the esterase for enantioselective outcomes).