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Pcal_1127, a highly stable and efficient ribose-5-phosphate pyrophosphokinase from Pyrobaculum calidifontis

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Abstract

Analysis of the genome sequence of Pyrobaculum calidifontis revealed the presence of an open reading frame Pcal_1127 annotated as ribose-5-phosphate pyrophosphokinase. To examine the properties of Pcal_1127 the coding gene was cloned, expressed in Escherichia coli, and the purified gene product was characterized. Pcal_1127 exhibited higher activity when ATP was replaced by dATP as pyrophosphate donor. Phosphate and EDTA activated the enzyme activity and equivalent amount of activity was detected with ATP and dATP in their presence. Recombinant Pcal_1127 could utilize all the four nucleotides as pyrophosphate donors with a marked preference for ATP. Optimum temperature and pH for the enzyme activity were 55 °C and 10.5, respectively. A unique feature of Pcal_1127 was its stability against temperature as well as denaturants. Pcal_1127 exhibited more than 95 % residual activity after heating for 4 h at 90 °C and a half-life of 15 min in the boiling water. The enzyme activity was not affected by the presence of 8 M urea or 4 M guanidinium chloride. Pcal_1127 was a highly efficient enzyme with a catalytic efficiency of 5183 mM−1 s−1. These features make Pcal_1127, a novel and unique ribose-5-phosphate pyrophosphokinase.

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Authors and Affiliations

  1. School of Biological Sciences, University of the Punjab, Quaid-e-Azam Campus, Lahore, 54590, Pakistan

    Tahira Bibi, Sumera Perveen, Iram Aziz, Qamar Bashir, Naeem Rashid & Muhammad Akhtar

  2. The Research Organization of Science and Technology, Ritsumeikan University, Kusatsu, Shiga, 525-8577, Japan

    Tadayuki Imanaka

  3. School of Biological Sciences, University of Southampton, Southampton, SO16 7PX, UK

    Muhammad Akhtar

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  1. Tahira Bibi
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  2. Sumera Perveen
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  3. Iram Aziz
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  4. Qamar Bashir
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  5. Naeem Rashid
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  6. Tadayuki Imanaka
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  7. Muhammad Akhtar
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Correspondence to Naeem Rashid.

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Communicated by H. Atomi.

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Bibi, T., Perveen, S., Aziz, I. et al. Pcal_1127, a highly stable and efficient ribose-5-phosphate pyrophosphokinase from Pyrobaculum calidifontis . Extremophiles 20, 821–830 (2016). https://doi.org/10.1007/s00792-016-0869-z

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  • DOI: https://doi.org/10.1007/s00792-016-0869-z

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