Poly‐3‐hydroxyalkanoate synthases from Pseudomonas putida U: substrate specificity and ultrastructural studies |
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Authors: | Sagrario Arias Angel Sandoval Mario Arcos Librada M Cañedo Beatriz Maestro Jesús M Sanz Germán Naharro José M Luengo |
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Institution: | 1. Departamento de Biología Molecular, Facultades de Veterinaria y de Biología, Universidad de León. 24071 León, Espa?a.;2. These authors have contributed equally to the development of this work.;3. Instituto Biomar, 24231 Onzonilla, León, Espa?a.;4. Instituto de Biología Molecular y Celular, Universidad Miguel Hernández, 03202 Elche (Alicante), Espa?a.;5. Departamento de Patología Animal (Sanidad Animal), Facultad de Veterinaria, Universidad de León. 24071 León, Espa?a. |
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Abstract: | The substrate specificity of the two polymerases (PhaC1 and PhaC2) involved in the biosynthesis of medium‐chain‐length poly‐hydroxyalkanoates (mcl PHAs) in Pseudomonas putida U has been studied in vivo. For these kind of experiments, two recombinant strains derived from a genetically engineered mutant in which the whole pha locus had been deleted (P. putida U Δpha) were employed. These bacteria, which expresses only phaC1 (P. putida U Δpha pMC‐phaC1) or only phaC2 (P. putida U Δpha pMC‐phaC2), accumulated different PHAs in function of the precursor supplemented to the culture broth. Thus, the P. putida U Δpha pMC‐phaC1 strain was able to synthesize several aliphatic and aromatic PHAs when hexanoic, heptanoic, octanoic decanoic, 5‐phenylvaleric, 6‐phenylhexanoic, 7‐phenylheptanoic, 8‐phenyloctanoic or 9‐phenylnonanoic acid were used as precursors; the highest accumulation of polymers was observed when the precursor used were decanoic acid (aliphatic PHAs) or 6‐phenylhexanoic acid (aromatic PHAs). However, although it synthesizes similar aliphatic PHAs (the highest accumulation was observed when hexanoic acid was the precursor) the other recombinant strain (P. putida U Δpha pMC‐phaC2) only accumulated aromatic PHAs when the monomer to be polymerized was 3‐hydroxy‐5‐phenylvaleryl‐CoA. The possible influence of the putative three‐dimensional structures on the different catalytic behaviour of PhaC1 and PhaC2 is discussed. |
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