title: Polyhydroyxalkanoate synthase fusions as a strategy for oriented enzyme immobilisation.
abstract: Polyhydroxyalkanoate (PHA) is a carbon storage polymer produced by certain bacteria in unbalanced nutrient conditions. The PHA forms spherical inclusions surrounded by granule associate proteins including the PHA synthase (PhaC). Recently, the intracellular formation of PHA granules with covalently attached synthase from Ralstonia eutropha has been exploited as a novel strategy for oriented enzyme immobilisation. Fusing the enzyme of interest to PHA synthase results in a bifunctional protein able to produce PHA granules and immobilise the active enzyme of choice to the granule surface. Functionalised PHA granules can be isolated from the bacterial hosts, such as Escherichia coli, and maintain enzymatic activity in a wide variety of assay conditions. This approach to oriented enzyme immobilisation has produced higher enzyme activities and product levels than non-oriented immobilisation techniques such as protein inclusion based particles. Here, enzyme immobilisation via PHA synthase fusion is reviewed in terms of the genetic designs, the choices of enzymes, the control of enzyme orientations, as well as their current and potential applications.
authors: Institute of Fundamental Sciences, Massey University, Private Bag 11222, Palmerston North 4442, New Zealand. d.o.hooks@massey.ac.nz.
authors: David O
authors: DO
authors: Hooks
authors: Institute of Fundamental Sciences, Massey University, Private Bag 11222, Palmerston North 4442, New Zealand. m.venningslater@massey.ac.nz.
authors: Mark
authors: M
authors: Venning-Slater
authors: Institute of Fundamental Sciences, Massey University, Private Bag 11222, Palmerston North 4442, New Zealand. j.du@massey.ac.nz.
authors: Jinping
authors: J
authors: Du
authors: Institute of Fundamental Sciences, Massey University, Private Bag 11222, Palmerston North 4442, New Zealand. b.rehm@massey.ac.nz.
authors: Bernd H A
authors: BH
authors: Rehm
mesh_terms: D000217:Acyltransferases / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000201:enzymology / Q000235:genetics; D020118:Cupriavidus necator / Q000201:enzymology; D004800:Enzymes, Immobilized / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D013294:Lactococcus lactis / Q000201:enzymology / Q000235:genetics; D009097:Multienzyme Complexes / Q000378:metabolism*; D054813:Polyhydroxyalkanoates / Q000096:biosynthesis; D011137:Polystyrenes; D020836:Protein Structure, Quaternary; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D004800:Enzymes, Immobilized; D009097:Multienzyme Complexes; D054813:Polyhydroxyalkanoates; D011137:Polystyrenes; D011993:Recombinant Fusion Proteins; C003771:Amberlite XAD-2 resin; D000217:Acyltransferases; C077225:poly(3-hydroxyalkanoic acid) synthase