Abstract
Polyhydroxyalkanoates (PHAs) are biopolymers synthesized by a wide range of bacteria, which serve as a promising candidate in replacing some conventional petrochemical-based plastics. PHA synthase (PhaC) is the key enzyme in the polymerization of PHA, and the crystal structures were successfully determined using the catalytic domain of PhaC from Cupriavidus necator (PhaCCn-CAT) and Chromobacterium sp. USM2 (PhaCCs-CAT). Here, we review the beneficial mutations discovered in PhaCs from a structural perspective. The structural comparison of the residues involved in beneficial mutation reveals that the residues are near to the catalytic triad, but not inside the catalytic pocket. For instance, Ala510 of PhaCCn is near catalytic His508 and may be involved in the open-close regulation, which presumably play an important role in substrate specificity and activity. In the class II PhaC1 from Pseudomonas sp. 61-3 (PhaC1Ps), Ser325 stabilizes the catalytic cysteine through hydrogen bonding. Another residue, Gln508 of PhaC1Ps is located in a conserved hydrophobic pocket which is next to the catalytic Asp and His. A class I, II-conserved Phe420 of PhaCCn is one of the residues involved in dimerization and its mutation to serine greatly reduced the lag phase. The current structural analysis shows that the Phe362 and Phe518 of PhaC from Aeromonas caviae (PhaCAc) are assisting the dimer formation and maintaining the integrity of the core beta-sheet, respectively. The structure-function relationship of PhaCs discussed in this review will serve as valuable reference for future protein engineering works to enhance the performance of PhaCs and to produce novel biopolymers.
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Funding
This work is supported by Research University Grant (RUI) from the Universiti Sains Malaysia (1001/PBIOLOGI/8011060 to K.S.), JST A-STEP Grant (29A1027 to T.H.) and JST CREST grant (JPMJCR12B4 to S.T.).
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Accession codes: atomic coordinates and structure factors used in this mini-review can be accessed from the Protein Data Bank (PDB) under ID codes: 5HZ2 (PhaCCn-CAT), 5T6O (PhaCCn-CAT), and 5XAV (PhaCCs-CAT).
The amino acid sequences used in the multiple sequence alignments are:Class I PhaCCs from Chromobacterium sp. USM2 (ADL70203), PhaCAc from Aeromonas caviae (BAA21815), PhaCCn from Cupriavidus necator (AAW65074);Class II PhaC1Ps from Pseudomonas sp. 61-3 (BAA36200), PhaC1Pa from Pseudomonas aeruginosa (BAQ42906);Class III PhaCAv from Allochromatium vinosum DSM180 (BAE20055), PhaCTv from Thiocystis violascens DSM198 (AFL75311);Class IV PhaCBm from Bacillus megaterium (AAD05260), PhaCBc from Bacillus cereus (BAI68395).
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Chek, M.F., Hiroe, A., Hakoshima, T. et al. PHA synthase (PhaC): interpreting the functions of bioplastic-producing enzyme from a structural perspective. Appl Microbiol Biotechnol 103, 1131–1141 (2019). https://doi.org/10.1007/s00253-018-9538-8
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DOI: https://doi.org/10.1007/s00253-018-9538-8