Structure of polyhydroxyalkanoate (PHA) synthase PhaC from Chromobacterium sp. USM2, producing biodegradable plastics

Abstract

Polyhydroxyalkanoate (PHA) is a promising candidate for use as an alternative bioplastic to replace petroleum-based plastics. Our understanding of PHA synthase PhaC is poor due to the paucity of available three-dimensional structural information. Here we present a high-resolution crystal structure of the catalytic domain of PhaC from Chromobacterium sp. USM2, PhaCCs-CAT. The structure shows that PhaCCs-CAT forms an α/β hydrolase fold comprising α/β core and CAP subdomains. The active site containing Cys291, Asp447 and His477 is located at the bottom of the cavity, which is filled with water molecules and is covered by the partly disordered CAP subdomain. We designated our structure as the closed form, which is distinct from the recently reported catalytic domain from Cupriavidus necator (PhaCCn-CAT). Structural comparison showed PhaCCn-CAT adopting a partially open form maintaining a narrow substrate access channel to the active site, but no product egress. PhaCCs-CAT forms a face-to-face dimer mediated by the CAP subdomains. This arrangement of the dimer is also distinct from that of the PhaCCn-CAT dimer. These findings suggest that the CAP subdomain should undergo a conformational change during catalytic activity that involves rearrangement of the dimer to facilitate substrate entry and product formation and egress from the active site.

Characterisation and modelling of a polyhydroxyalkanoate synthase fromAquitaleasp. USM4 reveals a mechanism for polymer elongation

“…Nevertheless, the present PhaC Cn structure is noted as partially open and could have been induced by the artificial disulphide bond between Cys382 located in the LID region and Cys438, none of which is conserved. This has led to part of its LID region (Leu358-Leu384) adopting an extended loop conformation encircling the opposite subunit, which in the closed-form PhaC Cs structure actually folded into an α-helix [8].…”

“…Recently the dimeric structures of class I PhaCs from Cupriavidus necator [6,7], PhaC Cn , and from Chromobacterium sp. USM2 [8], PhaC Cs , have been solved. The PhaC Cn structure is in a partially open form with an obvious channel proposed for substrate entrance, while the PhaC Cs structure is in a closed form with no visible access to the catalytic site.…”

Forest soil bacteria able to produce homo and copolymers of polyhydroxyalkanoates from several pure and waste carbon sources

“…Polyhydroxybutyrate (PHB) and poly(3‐hydroxybutyrate‐co‐3‐hydroxyvalerate) (PHB ‐co‐ HV) are the main PHAs representatives, which are produced from renewable sources and have similar properties to those of some petrochemical polymers such as polypropylene. Nowadays, the search of bacterial strains able to use a wide variety of carbon sources, including wastes, to produce PHAs (especially copolymers), is of great interest (Chek et al ). Copolymers of PHAs have different properties from those of PHB, like lower melting point, less crystallinity, more flexibility, higher viscosity in the liquid state and better mechanical properties, resulting in better processability and suitability for commercial exploitation (Rivera‐Briso and Serrano‐Aroca ).…”

Domain-centric dissection and classification of prokaryotic poly(3-hydroxyalkanoate) synthases

“…Except for the above-mentioned classical pathway, enzymes from several other pathways are also indirectly involved in PHA synthesis, such as 3-oxoacyl-[acyl-carrier-protein] reductase (FabG), (R)-specific enoyl-CoA hydratase (PhaJ), Malonyl CoA-acyl carrier protein transacylase (FabD), Succinate-CoA ligase [ADP-forming] subunit alpha (SucD), NAD-dependent 4-hydroxybutyrate dehydrogenase (4HbD), and 4-hydroxybutyrate-CoA:CoA transferase (OrzF), etc [4]. Currently, more than 150 different hydroxyalkanoic acids are known to occur as constituents of PHAs [6] and a total of 14 PHA synthesis pathways have been identified with many enzymes involved in these processes [7].…”

“…Class I, III and IV prefer SCL) carbon chain monomers (C3-5) while class II utilizes MCL PHA monomers (C6–14) [8]. In some exceptions, Class I PHA synthase could produce mixed PHAs incorporated with both SCL and MCL hydroxyalkanoates that show better functional properties compared with homo-polymers [2,7]. PhaC plays a key role in PHA synthesis and is the core subunit of PHA synthase.…”