Ayub Nicolás D, Julia Pettinari M, Méndez Beatriz S, López Nancy I
Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Buenos Aires, Argentina.
FEMS Microbiol Lett. 2006 Nov;264(1):125-31. doi: 10.1111/j.1574-6968.2006.00446.x.
Pseudomonas sp. 14-3 accumulates polyhydroxybutyrate (PHB) from octanoate, but not from glucose. To elucidate this unusual phenotype, genes responsible for the synthesis of PHB were cloned and analyzed. A PHB polymerase gene (phaC) was found downstream from genes coding for a beta-ketothiolase (phaA), an acetoacetyl-coenzyme A reductase (phaB) and a putative transcriptional regulator (phaR). All genes were similar to pha genes from several related species, but differences were observed in the distal region of phaA. Complementation with heterologous beta-ketothiolase genes from Azotobacter sp. FA8 or Pseudomonas putida GPp104 restored the capability of Pseudomonas sp. 14-3 to synthesize PHB from glucose, demonstrating that its beta-ketothiolase was nonfunctional. Analysis of the genome sequences of other Pseudomonas species has revealed the existence of putative beta-ketothiolase genes. The functionality of one of these thiolase genes, belonging to P. putida GPp104, was experimentally demonstrated. Pseudomonas sp. 14-3 is the first natural phaA mutant described, that despite this mutation accumulates high amounts of PHB when growing on fatty acids.
假单胞菌属14 - 3菌株能利用辛酸积累聚羟基丁酸酯（PHB），但不能利用葡萄糖。为阐明这种不寻常的表型，对负责PHB合成的基因进行了克隆和分析。在编码β - 酮硫解酶（phaA）、乙酰乙酰辅酶A还原酶（phaB）和一个假定的转录调节因子（phaR）的基因下游发现了一个PHB聚合酶基因（phaC）。所有基因都与几个相关物种的pha基因相似，但在phaA的远端区域观察到差异。用来自固氮菌属FA8或恶臭假单胞菌GPp104的异源β - 酮硫解酶基因进行互补，恢复了假单胞菌属14 - 3菌株从葡萄糖合成PHB的能力，表明其β - 酮硫解酶无功能。对其他假单胞菌物种的基因组序列分析揭示了假定的β - 酮硫解酶基因的存在。实验证明了其中一个属于恶臭假单胞菌GPp104的硫解酶基因的功能。假单胞菌属14 - 3菌株是描述的第一个天然phaA突变体，尽管有这种突变，但在脂肪酸上生长时仍能积累大量的PHB。
