Abstract
BackgroundPolyhydroxyalkanoates (PHA), are biodegradable polyesters derived from many microorganisms such as the pseudomonads. These polyesters are in great demand especially in the packaging industries, the medical line as well as the paint industries. The enzyme responsible in catalyzing the formation of PHA is PHA synthase. Due to the limited structural information, its functional properties including catalysis are lacking. Therefore, this study seeks to investigate the structural properties as well as its catalytic mechanism by predicting the three-dimensional (3D) model of the Type II Pseudomonas sp. USM 4–55 PHA synthase 1 (PhaC1P.sp USM 4–55).ResultsSequence analysis demonstrated that PhaC1P.sp USM 4–55 lacked similarity with all known structures in databases. PSI-BLAST and HMM Superfamily analyses demonstrated that this enzyme belongs to the alpha/beta hydrolase fold family. Threading approach revealed that the most suitable template to use was the human gastric lipase (PDB ID: 1HLG). The superimposition of the predicted PhaC1P.sp USM 4–55 model with 1HLG covering 86.2% of the backbone atoms showed an RMSD of 1.15 Å. The catalytic residues comprising of Cys296, Asp451 and His479 were found to be conserved and located adjacent to each other. In addition to this, an extension to the catalytic mechanism was also proposed whereby two tetrahedral intermediates were believed to form during the PHA biosynthesis. These transition state intermediates were further postulated to be stabilized by the formation of oxyanion holes. Based on the sequence analysis and the deduced model, Ser297 was postulated to contribute to the formation of the oxyanion hole.ConclusionThe 3D model of the core region of PhaC1P.sp USM 4–55 from residue 267 to residue 484 was developed using computational techniques and the locations of the catalytic residues were identified. Results from this study for the first time highlighted Ser297 potentially playing an important role in the enzyme's catalytic mechanism.
Highlights
Polyhydroxyalkanoates (PHA), are biodegradable polyesters derived from many microorganisms such as the pseudomonads
As the level of identity is very low, the pair-wise alignment described in Figure 1, even if it is suggestive of a potential relationship between the two sequences, may be inaccurate in some regions, this has been overcome by performing secondary structure prediction, multiple sequence alignment and threading procedures which will be further discussed below
A series of molecular modeling and computational methods were combined in order to gain insight into the 3D structure of PhaC1P.sp USM 4–55 concentrating on the α/β hydrolase fold region
Summary
Polyhydroxyalkanoates (PHA), are biodegradable polyesters derived from many microorganisms such as the pseudomonads. Polyhydroxyalkanoic acids (PHA) represent a complex class of biodegradable and naturally occurring biopolyesters that consist of hydroxyalkanoic acid monomers. They are produced by a wide range of bacteria as energy storage compounds especially during limited nutritional supplies and in the presence of excess carbon source. PHA synthase is the key enzyme that plays the central catalytic role in PHA production. It uses coenzymeA (CoA) thioesters of hydroxyalkanoic acids (HAs) as the main substrates and catalyzes the polymerization of HAs to yield PHA with the concomitant release of CoA [1,2]. They can be distinguished into four types based on the subunit composition and substrate specificities [6,7]
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