Cloning, Expression, and Characterization of the First Archaeal ATP-Dependent Glucokinase from Aerobic Hyperthermophilic Archaeon Aeropyrum pernix

2003 ◽  
Vol 133 (2) ◽  
pp. 219-224 ◽  
Author(s):  
H. Sakuraba
Keyword(s):  
Hyperthermophilic Archaeon ◽  
Aeropyrum Pernix

Purification and characterization of carbon monoxide dehydrogenase from the aerobic hyperthermophilic archaeon Aeropyrum pernix

2010 ◽  
Vol 76 (6) ◽  
pp. 999-1006 ◽  
Author(s):  
Hiroshi Nishimura ◽  
Yoshiko Nomura ◽  
Eri Iwata ◽  
Nozomi Sato ◽  
Yoshihiko Sako
Keyword(s):  
Carbon Monoxide ◽  
Carbon Monoxide Dehydrogenase ◽  
Purification And Characterization ◽  
Hyperthermophilic Archaeon ◽  
Aeropyrum Pernix

Characterization of native glutamate dehydrogenase from an aerobic hyperthermophilic archaeon Aeropyrum pernix K1

2001 ◽  
Vol 56 (3-4) ◽  
pp. 388-394 ◽  
Author(s):  
I. Helianti ◽  
Y. Morita ◽  
A. Yamamura ◽  
Y. Murakami ◽  
K. Yokoyama ◽  
...  
Keyword(s):  
Glutamate Dehydrogenase ◽  
Hyperthermophilic Archaeon ◽  
Aeropyrum Pernix

scholarly journals Characterization of a Novel Thermostable O-Acetylserine Sulfhydrylase from Aeropyrum pernix K1

2003 ◽  
Vol 185 (7) ◽  
pp. 2277-2284 ◽  
Author(s):  
Koshiki Mino ◽  
Kazuhiko Ishikawa
Keyword(s):  
Escherichia Coli ◽  
Rate Constant ◽  
Binding Motif ◽  
Phosphate Binding ◽  
Ph Optimum ◽  
Hyperthermophilic Archaeon ◽  
Aeropyrum Pernix ◽  
Synthetic Reaction

ABSTRACT An O-acetylserine sulfhydrylase (OASS) from the hyperthermophilic archaeon Aeropyrum pernix K1, which shares the pyridoxal 5′-phosphate binding motif with both OASS and cystathionine β-synthase (CBS), was cloned and expressed by using Escherichia coli Rosetta(DE3). The purified protein was a dimer and contained pyridoxal 5′-phosphate. It was shown to be an enzyme with CBS activity as well as OASS activity in vitro. The enzyme retained 90% of its activity after a 6-h incubation at 100°C. In the O-acetyl-l-serine sulfhydrylation reaction, it had a pH optimum of 6.7, apparent Km values for O-acetyl-l-serine and sulfide of 28 and below 0.2 mM, respectively, and a rate constant of 202 s−1. In the l-cystathionine synthetic reaction, it showed a broad pH optimum in the range of 8.1 to 8.8, apparent Km values for l-serine and l-homocysteine of 8 and 0.51 mM, respectively, and a rate constant of 0.7 s−1. A. pernix OASS has a high activity in the l-cysteine desulfurization reaction, which produces sulfide and S-(2,3-hydroxy-4-thiobutyl)-l-cysteine from l-cysteine and dithiothreitol.

scholarly journals Gene Expression and Characterization of a Third Type of Dye-LinkedL-Proline Dehydrogenase from the Aerobic Hyperthermophilic Archaeon,Aeropyrum pernix

2012 ◽  
Vol 76 (3) ◽  
pp. 589-593 ◽  
Author(s):  
Takenori SATOMURA ◽  
Yusuke HARA ◽  
Shin-ichiro SUYE ◽  
Haruhiko SAKURABA ◽  
Toshihisa OHSHIMA
Keyword(s):  
Gene Expression ◽  
Proline Dehydrogenase ◽  
Hyperthermophilic Archaeon ◽  
Aeropyrum Pernix

Characterization of a novel moderate-substrate specificity amino acid racemase from the hyperthermophilic archaeon Thermococcus litoralis

2021 ◽  
Author(s):  
Ryushi Kawakami ◽  
Chinatsu Kinoshita ◽  
Tomoki Kawase ◽  
Mikio Sato ◽  
Junji Hayashi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Substrate Specificity ◽  
Enzyme Stability ◽  
Hyperthermophilic Archaea ◽  
Thermococcus Litoralis ◽  
Hyperthermophilic Archaeon ◽  
Aminobutyrate Aminotransferase ◽  
Chaperone Protein ◽  
Amino Acid Racemase

Abstract The amino acid sequence of the OCC_10945 gene product from the hyperthermophilic archaeon Thermococcus litoralis DSM5473, originally annotated as γ-aminobutyrate aminotransferase, is highly similar to that of the uncharacterized pyridoxal 5ʹ-phosphate (PLP)-dependent amino acid racemase from Pyrococcus horikoshii. The OCC_10945 enzyme was successfully overexpressed in Escherichia coli by co-expression with a chaperone protein. The purified enzyme demonstrated PLP-dependent amino acid racemase activity primarily toward Met and Leu. Although PLP contributed to enzyme stability, it only loosely bound to this enzyme. Enzyme activity was strongly inhibited by several metal ions, including Co2+ and Zn2+, and non-substrate amino acids such as l-Arg and l-Lys. These results suggest that the underlying PLP-binding and substrate recognition mechanisms in this enzyme are significantly different from those of the other archaeal and bacterial amino acid racemases. This is the first description of a novel PLP-dependent amino acid racemase with moderate substrate specificity in hyperthermophilic archaea.

scholarly journals First Characterization of an Archaeal GTP-Dependent Phosphoenolpyruvate Carboxykinase from the Hyperthermophilic Archaeon Thermococcus kodakaraensis KOD1

2004 ◽  
Vol 186 (14) ◽  
pp. 4620-4627 ◽  
Author(s):  
Wakao Fukuda ◽  
Toshiaki Fukui ◽  
Haruyuki Atomi ◽  
Tadayuki Imanaka
Keyword(s):  
Binding Sites ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Cation Binding ◽  
Activity Levels ◽  
Binding Region ◽  
Hyperthermophilic Archaeon ◽  
Dependent Activity ◽  
Additional Role

ABSTRACT Phosphoenolpyruvate carboxykinase (PCK), which catalyzes the nucleotide-dependent, reversible decarboxylation of oxaloacetate to yield phosphoenolpyruvate and CO2, is one of the important enzymes in the interconversion between C3 and C4 metabolites. This study focused on the first characterization of the enzymatic properties and expression profile of an archaeal PCK from the hyperthermophilic archaeon Thermococcus kodakaraensis (Pck Tk ). Pck Tk showed 30 to 35% identities to GTP-dependent PCKs from mammals and bacteria but was located in a branch distinct from that of the classical enzymes in the phylogenetic tree, together with other archaeal homologs from Pyrococcus and Sulfolobus spp. Several catalytically important regions and residues, found in all known PCKs irrespective of their nucleotide specificities, were conserved in Pck Tk . However, the predicted GTP-binding region was unique compared to those in other GTP-dependent PCKs. The recombinant Pck Tk actually exhibited GTP-dependent activity and was suggested to possess dual cation-binding sites specific for Mn2+ and Mg2+. The enzyme preferred phosphoenolpyruvate formation from oxaloacetate, since the Km value for oxaloacetate was much lower than that for phosphoenolpyruvate. The transcription and activity levels in T. kodakaraensis were higher under gluconeogenic conditions than under glycolytic conditions. These results agreed with the role of Pck Tk in providing phosphoenolpyruvate from oxaloacetate as the first step of gluconeogenesis in this hyperthermophilic archaeon. Additionally, under gluconeogenic conditions, we observed higher expression levels of Pck Tk on pyruvate than on amino acids, implying that it plays an additional role in the recycling of excess phosphoenolpyruvate produced from pyruvate, replacing the function of the anaplerotic phosphoenolpyruvate carboxylase that is missing from this archaeon.

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