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Purification and functional characterization of thermostable 5-aminolevulinic acid synthases

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Abstract

Objectives

As 5-aminolevulinic acid synthase (ALAS), the key enzyme for 5-aminolevulinic acid (ALA) synthesis, is unstable, we have sought to find thermostable ALASs from thermophilic organisms.

Results

Three ALASs from thermophiles Geobacillus thermoglucosidasius (GT-ALAS), Laceyella sacchari (LS-ALAS) and Pseudomonas alcaliphila (PA-ALAS) were purified and characterized. All enzymes were more stable than two previously studied ALASs from Rhodopseudomonas palustris and Rhodobacter sphaeroides. There was almost no activity change after 60 h at 37 °C for the three thermostable enzymes. This contrasts with the other two enzymes which lost over 90 % activities in just 1 h. Furthermore, the specific activity of LS-ALAS (7.8 U mg−1) was also higher than any previously studied ALASs.

Conclusions

Thermostable ALASs were found in thermophilic organisms and this paves the way for developing cell free processes for enzymatic production of ALA from bulk chemicals succinate and glycine.

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Acknowledgments

This work was supported by the National Key Basic Research Program of China (973 Program) (Nos. 2012CB725203, 2011CBA00804), National High Technology Research and Development Program of China (863 Program) (No. 2012AA022103) and Natural Science Foundation of Tianjin (No. 12JCYBJC33000).

Supporting information

Supplementary Table 1—Bacterial strains and plasmids used in this study.

Supplementary Table 2—The NCBI protein IDs of the studied enzymes.

Supplementary Fig. 1—Comparison of amino acid sequences of ALASs from different organisms.

Supplementary Fig. 2—Initial enzymatic reaction rates measured at various substrate concentrations.

Supplementary Fig. 3—Influence of PLP addition during the purification process on the specific activities of ALASs.

Author information

Authors and Affiliations

  1. School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, People’s Republic of China

    Qinglong Meng, Xueming Zhao & Tao Chen

  2. Key Laboratory of Systems Microbial Biotechnology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 West 7th Avenue, Tianjin Airport Economic Area, Tianjin, 300308, People’s Republic of China

    Qinglong Meng, Yanfei Zhang, Chunling Ma & Hongwu Ma

Authors
  1. Qinglong Meng
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  2. Yanfei Zhang
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  3. Chunling Ma
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  4. Hongwu Ma
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  5. Xueming Zhao
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  6. Tao Chen
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Corresponding author

Correspondence to Hongwu Ma.

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Meng, Q., Zhang, Y., Ma, C. et al. Purification and functional characterization of thermostable 5-aminolevulinic acid synthases. Biotechnol Lett 37, 2247–2253 (2015). https://doi.org/10.1007/s10529-015-1903-4

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  • DOI: https://doi.org/10.1007/s10529-015-1903-4

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