Two different pathways are involved in the β‐oxidation of n‐alkanoic and n‐phenylalkanoic acids in Pseudomonas putida U: genetic studies and biotechnological applications

Abstract

In Pseudomonas putida U, the degradation of n‐alkanoic and n‐phenylalkanoic acids is carried out by two sets of β‐oxidation enzymes (βI and βII). Whereas the first one (called βI) is constitutive and catalyses the degradation of n‐alkanoic and n‐phenylalkanoic acids very efficiently, the other one (βII), which is only expressed when some of the genes encoding βI enzymes are mutated, catabolizes n‐phenylalkanoates (n > 4) much more slowly. Genetic studies revealed that disruption or deletion of some of the βI genes handicaps the growth of P. putida U in media containing n‐alkanoic or n‐phenylalkanoic acids with an acyl moiety longer than C4. However, all these mutants regained their ability to grow in media containing n‐alkanoates as a result of the induction of βII, but they were still unable to catabolize n‐phenylalkanoates completely, as the βI‐FadBA enzymes are essential for the β‐oxidation of certain n‐phenylalkanoyl‐CoA derivatives when they reach a critical size. Owing to the existence of the βII system, mutants lacking βIfadB/A are able to synthesize new poly 3‐OH‐n‐alkanoates (PHAs) and poly 3‐OH‐n‐phenylalkanoates (PHPhAs) efficiently. However, they are unable to degrade these polymers, becoming bioplastic overproducer mutants. The genetic and biochemical importance of these results is reported and discussed.

Making Green Polymers Even Greener:Towards Sustainable Production of Polyhydroxyalkanoates from Agroindustrial By-Products

“…Molecular biology strategies designed to increase the production of MCL-PHA in Pseudomonas was firstly described in P. putida U (García et al, 1999). The existence in the genome of this strain of several sets of iso-enzymes encoding genes similar to those belonging to the fad regulon from E. coli from the -oxidation of fatty acids have been described (Olivera et al, 2001a(Olivera et al, , 2001b. Engineered strains carrying mutations in the fadA-fadB genes had a strong intracellular accumulation of biopolyesters.…”

Plasmids containing the same origin of replication are useful tools to perform biotechnological studies in Pseudomonas putida U and in E. coli DH10B

“…P. putida fadBA (PpU fadBA) is a poly-3-hydroxy-n-alkanoate (PHAs) overproducer mutant in which the fadBA genes, encoding the proteins FadB and FadA belonging to the main β-oxidation pathway, have been deleted [8]. This mutant, although slowly, grows in minimal media containing n-alkanoic acids as the sole carbon source, but it does not grow in media containing n-aryl-alkanoic acids.…”

Nitrile Pathway Involving Acyl-CoA Synthetase

“…The protein encoded by acsA showed the highest amino acid sequence similarity with FadD1 (70% identity) from Pseudomonas putida U (Ref. 42; GenBank TM accession no AF290950), whose mutant is unable to grow in a minimum medium containing an n-alkanoic acid or n-phenylalkanoic acid (with an acyl chain of longer than C4) as sole carbon source. AcsA is homologous to acetyl-CoA synthetase (31% identity) and acyl-CoA synthetase (25% identity) from E. coli (GenBank TM accession numbers U00006 and L02649, respectively).…”