Abstract
BackgroundThe genus Thermus, which has been considered for a long time as a fruitful source of biotechnological relevant enzymes, has emerged more recently as suitable host to overproduce thermozymes. Among these, α-galactosidases are widely used in several industrial bioprocesses that require high working temperatures and for which thermostable variants offer considerable advantages over their thermolabile counterparts.ResultsThermus thermophilus HB27 strain was used for the homologous expression of the TTP0072 gene encoding for an α-galactosidase (TtGalA). Interestingly, a soluble and active histidine-tagged enzyme was produced in larger amounts (5 mg/L) in this thermophilic host than in Escherichia coli (0.5 mg/L). The purified recombinant enzyme showed an optimal activity at 90 °C and retained more than 40% of activity over a broad range of pH (from 5 to 8).ConclusionsTtGalA is among the most thermoactive and thermostable α-galactosidases discovered so far, thus pointing to T. thermophilus as cell factory for the recombinant production of biocatalysts active at temperature values over 90 °C.
Highlights
The genus Thermus, which has been considered for a long time as a fruitful source of biotechnological relevant enzymes, has emerged more recently as suitable host to overproduce thermozymes
The only protein sequence retrieved is encoded by TTP0072 (Accession No AAS82402) and shows high sequence identity with previously characterized α-galactosidases from Thermus sp. strain T2 (75%; Accession No BAA76597), Thermus brockianus (73%; Accession No AF135398), Thermotoga neapolitana (36%; Accession No AF011400), Thermotoga maritima (35%; Accession No AJ001776), Sulfolobus solfataricus (39%; Accession No Q97U94) and H. sapiens (23%; Accession No CAA29232.1)
A multiple alignment (Fig. 1a) revealed the presence of a consensus motif (FEVFQIDDGW) in the TTP0072 translated sequence characteristic of α-galactosidases. This is generally localised within the central region of bacterial enzymes (CAZy family GH36) or at the amino-terminal region of eukaryotic variants (CAZy family GH27)
Summary
The genus Thermus, which has been considered for a long time as a fruitful source of biotechnological relevant enzymes, has emerged more recently as suitable host to overproduce thermozymes. Mannans comprise linear or branched polymers derived from sugars such as d-mannose, d-galactose and d-glucose. They represent the major source of secondary cell wall found in conifers (softwood) and leguminosae [2]. Based on their sugar composition, mannans are classified in four α-Galactosidases (α-d-galactoside galactohydrolase EC 3.2.1.22) are exoglycosidases that catalyse the cleavage of the terminal non-reducing α-1,6-linked galactose residues present in different galactose-containing oligo- and polysaccharides. Α-galactosidases offer a promising solution for the degradation of raffinose in beet sugar industry [10], pulp and paper manufacturing [11] as well as in soy food [12] and animal feed processing [13]
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