caa a22 4500 445818751 CHVBK 20180317145237.0 cr unu---uuuuu 170323e20110301xx s 000 0 eng 10.1007/s00792-011-0355-6 doi (NATIONALLICENCE)springer-10.1007/s00792-011-0355-6 Gene cloning and protein expression of γ-glutamyltranspeptidases from Thermus thermophilus and Deinococcus radiodurans [Elektronische Daten] comparison of molecular and structural properties with mesophilic counterparts [Immacolata Castellano, Anna Di Salle, Antonello Merlino, Mosè Rossi, Francesco La Cara] γ-Glutamyltranspeptidase (γ-GT) is an ubiquitous enzyme that catalyzes the hydrolysis of γ-glutamyl bonds in glutathione and glutamine and the transfer of the released γ-glutamyl group to amino acids or short peptides. γ-GTs from extremophiles, bacteria adapted to live in hostile environments, were selected as model systems to study the molecular underpinnings of their adaptation to extreme conditions and to find out special properties of potential biotechnological interest. Here, we report the cloning, expression and purification of two members of γ-GT family from two different extremophilic species, Thermus thermophilus (TtGT) and Deinococcus radiodurans (DrGT); the first is an aerobic eubacterium, growing at high temperatures (50-82°C), the second is a polyextremophile, as it tolerates radiations, cold, dehydration, vacuum, and acid. TtGT and DrGT were both synthesized as precursor proteins of 59-60kDa, undergoing an intramolecular auto-cleavage to yield two subunits of 40 and 19-20kDa, respectively. However, like the γ-GT from Geobacillus thermodenitrificans, but differently from the other characterized bacterial and eukaryotic γ-GTs, the two new extremophilic enzymes displayed γ-glutamyl hydrolase, but not transpeptidase activity in the 37-50°C temperature range, pH 8.0. The comparison of sequences and structural models of these two proteins with experimental-determined structures of other known mesophilic γ-GTs suggests that the extremophilic members of this protein family have found a common strategy to adapt to different hostile environments. Moreover, a phylogenetic analysis suggests that γ-GTs displaying only γ-glutamyl hydrolase activity could represent the progenitors of the bacterial and eukaryotic counterparts. Springer, 2011 Gamma-glutamyltranspeptidases nationallicence Molecular adaptation nationallicence Extremophiles nationallicence Thermus thermophilus nationallicence Deinococcus radiodurans nationallicence γ-GT : Gamma-glutamyltranspeptidase nationallicence Bs : Bacillus subtilis nationallicence Bh : Bacillus halodurans nationallicence Dr : Deinococcus radiodurans nationallicence Ec : Escherichia coli nationallicence Gth : Geobacillus thermodenitrificans nationallicence Hp : Helicobacter pylori nationallicence h : human nationallicence Ta : Thermoplasma acidophilum nationallicence Tt : Thermus thermophilus nationallicence GNA : l-Glutamic acid γ-(4-nitroanilide) nationallicence GSH : Glutathione nationallicence Castellano Immacolata CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut Di Salle Anna CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut Merlino Antonello Department of Chemistry, University of Naples ‘Federico II', via Cintia, 80126, Naples, Italy aut Rossi Mosè CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut La Cara Francesco CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut Extremophiles Springer Japan 15/2(2011-03-01), 259-270 1431-0651 15:2<259 2011 15 792 https://doi.org/10.1007/s00792-011-0355-6 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00792-011-0355-6 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Castellano Immacolata CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut NATIONALLICENCE 700 1- Di Salle Anna CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut NATIONALLICENCE 700 1- Merlino Antonello Department of Chemistry, University of Naples ‘Federico II', via Cintia, 80126, Naples, Italy aut NATIONALLICENCE 700 1- Rossi Mosè CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut NATIONALLICENCE 700 1- La Cara Francesco CNR, Institute of Protein Biochemistry, via Pietro Castellino 111, 80131, Naples, Italy aut NATIONALLICENCE 773 0- Extremophiles Springer Japan 15/2(2011-03-01), 259-270 1431-0651 15:2<259 2011 15 792 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer