Displaying publications 1 - 20 of 47 in total

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  1. N-terminal truncation of PhaCBP-M-CPF4 and its effect on PHA production
    Neoh SZ, Tan HT, Trakunjae C, Chek MF, Vaithanomsat P, Hakoshima T, et al.
    Microb Cell Fact, 2024 Feb 15;23(1):52.
    PMID: 38360657 DOI: 10.1186/s12934-024-02329-w
    BACKGROUND: Among the polyhydroxyalkanoate (PHA), poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)] is reported to closely resemble polypropylene and low-density polyethylene. Studies have shown that PHA synthase (PhaC) from mangrove soil (PhaCBP-M-CPF4) is an efficient PhaC for P(3HB-co-3HHx) production and N-termini of PhaCs influence its substrate specificity, dimerization, granule morphology, and molecular weights of PHA produced. This study aims to further improve PhaCBP-M-CPF4 through N-terminal truncation.

    RESULTS: The N-terminal truncated mutants of PhaCBP-M-CPF4 were constructed based on the information of the predicted secondary and tertiary structures using PSIPRED server and AlphaFold2 program, respectively. The N-terminal truncated PhaCBP-M-CPF4 mutants were evaluated in C. necator mutant PHB-4 based on the cell dry weight, PHA content, 3HHx molar composition, molecular weights, and granule morphology of the PHA granules. The results showed that most transformants harbouring the N-terminal truncated PhaCBP-M-CPF4 showed a reduction in PHA content and cell dry weight except for PhaCBP-M-CPF4 G8. PhaCBP-M-CPF4 G8 and A27 showed an improved weight-average molecular weight (Mw) of PHA produced due to lower expression of the truncated PhaCBP-M-CPF4. Transformants harbouring PhaCBP-M-CPF4 G8, A27, and T74 showed a reduction in the number of granules. PhaCBP-M-CPF4 G8 produced higher Mw PHA in mostly single larger PHA granules with comparable production as the full-length PhaCBP-M-CPF4.

    CONCLUSION: This research showed that N-terminal truncation had effects on PHA accumulation, substrate specificity, Mw, and granule morphology. This study also showed that N-terminal truncation of the amino acids that did not adopt any secondary structure can be an alternative to improve PhaCs for the production of PHA with higher Mw in mostly single larger granules.

    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism
  2. Complete genome sequence of Aquitalea pelogenes USM4 (JCM19919), a polyhydroxyalkanoate producer
    Wan JH, Ng LM, Neoh SZ, Kajitani R, Itoh T, Kajiwara S, et al.
    Arch Microbiol, 2023 Jan 16;205(2):66.
    PMID: 36645481 DOI: 10.1007/s00203-023-03406-1
    Polyhydroxyalkanoate (PHA) is a type of biopolymer produced by most bacteria and archaea, resembling thermoplastic with biodegradability and biocompatibility features. Here, we report the complete genome of a PHA producer, Aquitalea sp. USM4, isolated from Perak, Malaysia. This bacterium possessed a 4.2 Mb circular chromosome and a 54,370 bp plasmid. A total of 4067 predicted protein-coding sequences, 87 tRNA genes, and 25 rRNA operons were identified using PGAP. Based on ANI and dDDH analysis, the Aquitalea sp. USM4 is highly similar to Aquitalea pelogenes. We also identified genes, including acetyl-CoA (phaA), acetoacetyl-CoA (phaB), PHA synthase (phaC), enoyl-CoA hydratase (phaJ), and phasin (phaP), which play an important role in PHA production in Aquitalea sp. USM4. The heterologous expression of phaC1 from Aquitalea sp. USM4 in Cupriavidus necator PHB-4 was able to incorporate six different types of PHA monomers, which are 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), 4-hydroxybutyrate (4HB), 5-hydroxyvalerate (5HV), 3-hydroxyhexanoate (3HHx) and isocaproic acid (3H4MV) with suitable precursor substrates. This is the first complete genome sequence of the genus Aquitalea among the 22 genome sequences from 4 Aquitalea species listed in the GOLD database, which provides an insight into its genome evolution and molecular machinery responsible for PHA biosynthesis.
    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism
  3. Identification of regions affecting enzyme activity, substrate binding, dimer stabilization and polyhydroxyalkanoate (PHA) granule morphology in the PHA synthase of Aquitalea sp. USM4
    Lim H, Chuah JA, Chek MF, Tan HT, Hakoshima T, Sudesh K
    Int J Biol Macromol, 2021 Sep 01;186:414-423.
    PMID: 34246679 DOI: 10.1016/j.ijbiomac.2021.07.041
    Polyhydroxyalkanoates (PHAs) are biopolyesters synthesized by microorganisms as intracellular energy reservoirs under stressful environmental conditions. PHA synthase (PhaC) is the key enzyme responsible for PHA biosynthesis, but the importance of its N- and C-terminal ends still remains elusive. Six plasmid constructs expressing truncation variants of Aquitalea sp. USM4 PhaC (PhaC1As) were generated and heterologously expressed in Cupriavidus necator PHB-4. Removal of the first six residues at the N-terminus enabled the modulation of PHA composition without altering the PHA content in cells. Meanwhile, deletion of 13 amino acids from the C-terminus greatly affected the catalytic activity of PhaC1As, retaining only 1.1-7.4% of the total activity. Truncation(s) at the N- and/or C-terminus of PhaC1As gradually diminished the incorporation of comonomer units, and revealed that the N-terminal region is essential for PhaC1As dimerization whereas the C-terminal region is required for stabilization. Notably, transmission electron microscopy analysis showed that PhaC modification affected the morphology of intracellular PHA granules, which until now is only known to be regulated by phasins. This study provided substantial evidence and highlighted the significance of both the N- and C-termini of PhaC1As in regulating intracellular granule morphology, activity, substrate specificity, dimerization and stability of the synthase.
    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism*; Acyltransferases/chemistry
  4. Evaluation of BP-M-CPF4 polyhydroxyalkanoate (PHA) synthase on the production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) from plant oil using Cupriavidus necator transformants
    Tan HT, Chek MF, Lakshmanan M, Foong CP, Hakoshima T, Sudesh K
    Int J Biol Macromol, 2020 Sep 15;159:250-257.
    PMID: 32417540 DOI: 10.1016/j.ijbiomac.2020.05.064
    Among the various types of polyhydroxyalkanoate (PHA), poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)] has a high potential to serve as commercial bioplastic due to its striking resemblance to petroleum-based plastics. In this study, five different genotypes of Cupriavidusnecator transformants harbouring the phaCBP-M-CPF4 gene (including PHB¯4/pBBR1-CBP-M-CPF4) were developed to evaluate the efficiency of 3HHx monomer incorporation. The fraction of 3-hydroxyhexanoate (3HHx) monomer that was incorporated into the PHA synthesized by these C. necator transformants using palm oil as the sole carbon source, was examined. Overall, co-expression of enoyl-CoA hydratase gene (phaJ1) from Pseudomonas aeruginosa, along with PHA synthase (PhaC), increased the 3HHx composition in the PHA copolymer. The differences in the enzyme activities of β-ketothiolase (PhaACn) and NADPH-dependent acetoacetyl-CoA reductase (PhaBCn) of the C. necator mutant hosts used in this study, were observed to alter the 3HHx composition and molecular weight of the PHA copolymer produced. The 3HHx fractions in the P(3HB-co-3HHx) produced by these C. necator transformants ranged between 1 and 18 mol%, while the weight-average molecular weight ranged from 0.7 × 106 to 1.8 × 106 Da. PhaCBP-M-CPF4 displayed a typical initial lag-phase and a relatively low synthase activity in the in vitro enzyme assay, which is thought to be the reason for the higher molecular weights of PHA obtained in this study.
    Matched MeSH terms: Acyltransferases/metabolism*
  5. Fulltext Asymmetric Open-Closed Dimer Mechanism of Polyhydroxyalkanoate Synthase PhaC
    Chek MF, Kim SY, Mori T, Tan HT, Sudesh K, Hakoshima T
    iScience, 2020 May 22;23(5):101084.
    PMID: 32388399 DOI: 10.1016/j.isci.2020.101084
    Biodegradable polyester polyhydroxyalkanoate (PHA) is a promising bioplastic material for industrial use as a replacement for petroleum-based plastics. PHA synthase PhaC forms an active dimer to polymerize acyl moieties from the substrate acyl-coenzyme A (CoA) into PHA polymers. Here we present the crystal structure of the catalytic domain of PhaC from Chromobacterium sp. USM2, bound to CoA. The structure reveals an asymmetric dimer, in which one protomer adopts an open conformation bound to CoA, whereas the other adopts a closed conformation in a CoA-free form. The open conformation is stabilized by the asymmetric dimerization and enables PhaC to accommodate CoA and also to create the product egress path. The bound CoA molecule has its β-mercaptoethanolamine moiety extended into the active site with the terminal SH group close to active center Cys291, enabling formation of the reaction intermediate by acylation of Cys291.
    Matched MeSH terms: Acyltransferases
  6. Polyhydroxyalkanoate (PHA) synthase genes and PHA-associated gene clusters in Pseudomonas spp. and Janthinobacterium spp. isolated from Antarctica
    Tan IKP, Foong CP, Tan HT, Lim H, Zain NA, Tan YC, et al.
    J Biotechnol, 2020 Apr 10;313:18-28.
    PMID: 32171790 DOI: 10.1016/j.jbiotec.2020.03.006
    The polyhydroxyalkanoate (PHA) producing capability of four bacterial strains isolated from Antarctica was reported in a previous study. This study analyzed the PHA synthase genes and the PHA-associated gene clusters from the two antarctic Pseudomonas isolates (UMAB-08 and UMAB-40) and the two antarctic Janthinobacterium isolates (UMAB-56 and UMAB-60) through whole-genome sequence analysis. The Pseudomonas isolates were found to carry PHA synthase genes which fall into two different PHA gene clusters, namely Class I and Class II, which are involved in the biosynthesis of short-chain-length-PHA (SCL-PHA) and medium-chain-length-PHA (MCL-PHA), respectively. On the other hand, the Janthinobacterium isolates carry a Class I and an uncharacterized putative PHA synthase genes. No other gene involved in PHA synthesis was detected in close proximity to the uncharacterized putative PHA synthase gene in the Janthinobacterium isolates, therefore it falls into a separate clade from the ordinary Class I, II, III and IV clades of PHA synthase (PhaC) phylogenetic tree. Multiple sequence alignment showed that the uncharacterized putative PHA synthase gene contains all the highly conserved amino acid residues and the proposed catalytic triad of PHA synthase. PHA biosynthesis and in vitro PhaC enzymatic assay results showed that this uncharacterized putative PHA synthase from Janthinobacterium sp. UMAB-60 is funtional. This report adds new knowledge to the PHA synthase database as we describe scarce information of PHA synthase genes and PHA-associated gene clusters from the antarctic bacterial isolates (extreme and geographically isolated environment) and comparing with those from non-antarctic PHA-producing bacteria.
    Matched MeSH terms: Acyltransferases
  7. Complete Genome Sequence of a Novel Polyhydroxyalkanoate (PHA) Producer, Jeongeupia sp. USM3 (JCM 19920) and Characterization of Its PHA Synthases
    Zain NA, Ng LM, Foong CP, Tai YT, Nanthini J, Sudesh K
    Curr Microbiol, 2020 Mar;77(3):500-508.
    PMID: 31893298 DOI: 10.1007/s00284-019-01852-z
    A novel polyhydroxyalkanoate (PHA)-producing bacterium, Jeongeupia sp. USM3 (JCM 19920) was isolated from the limestone soil at Gua Tempurung, Perak, Malaysia. This is the first report on the complete genome sequence for the genus Jeongeupia. This genome consists of a circular chromosome with a size of 3,788,814 bp and contains 3557 genes. Two PHA synthase (phaC) genes encoding for the key enzyme in the polymerization of PHA monomers and other PHA-associated genes were identified from the genome. Phylogenetic analysis of the PhaC protein sequences has revealed that both PhaC1 and PhaC2 of Jeongeupia sp. USM3 are categorized as Class I PHA synthases with 56% similarity to each other. Both of the PHA synthase genes of this isolate were cloned and heterologously expressed in a PHA mutant strain Cupriavidus necator PHB-4. The ability of the transformants to accumulate PHA showed that both PhaC1 and PhaC2 were functional.
    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism*
  8. Fulltext The chromosome-scale reference genome of black pepper provides insight into piperine biosynthesis
    Hu L, Xu Z, Wang M, Fan R, Yuan D, Wu B, et al.
    Nat Commun, 2019 10 16;10(1):4702.
    PMID: 31619678 DOI: 10.1038/s41467-019-12607-6
    Black pepper (Piper nigrum), dubbed the 'King of Spices' and 'Black Gold', is one of the most widely used spices. Here, we present its reference genome assembly by integrating PacBio, 10x Chromium, BioNano DLS optical mapping, and Hi-C mapping technologies. The 761.2 Mb sequences (45 scaffolds with an N50 of 29.8 Mb) are assembled into 26 pseudochromosomes. A phylogenomic analysis of representative plant genomes places magnoliids as sister to the monocots-eudicots clade and indicates that black pepper has diverged from the shared Laurales-Magnoliales lineage approximately 180 million years ago. Comparative genomic analyses reveal specific gene expansions in the glycosyltransferase, cytochrome P450, shikimate hydroxycinnamoyl transferase, lysine decarboxylase, and acyltransferase gene families. Comparative transcriptomic analyses disclose berry-specific upregulated expression in representative genes in each of these gene families. These data provide an evolutionary perspective and shed light on the metabolic processes relevant to the molecular basis of species-specific piperine biosynthesis.
    Matched MeSH terms: Acyltransferases/genetics
  9. Fulltext Transcriptional regulation of Lonicera japonica Thunb. during flower development as revealed by comprehensive analysis of transcription factors
    Wang T, Yang B, Guan Q, Chen X, Zhong Z, Huang W, et al.
    BMC Plant Biol, 2019 May 14;19(1):198.
    PMID: 31088368 DOI: 10.1186/s12870-019-1803-1
    BACKGROUND: Lonicera japonica Thunb. flower has been used for the treatment of various diseases for a long time and attracted many studies on its potential effects. Transcription factors (TFs) regulate extensive biological processes during plant development. As the restricted reports of L. japonica on TFs, our work was carried out to better understand the TFs' regulatory roles under different developmental stages in L. japonica.

    RESULTS: In this study, 1316 TFs belonging to 52 families were identified from the transcriptomic data, and corresponding expression profiles during the L. japonica flower development were comprehensively analyzed. 917 (69.68%) TFs were differentially expressed. TFs in bHLH, ERF, MYB, bZIP, and NAC families exhibited obviously altered expression during flower growth. Based on the analysis of differentially expressed TFs (DETFs), TFs in MYB, WRKY, NAC and LSD families that involved in phenylpropanoids biosynthesis, senescence processes and antioxidant activity were detected. The expression of MYB114 exhibited a positive correlation with the contents of luteoloside; Positive correlation was observed among the expression of MYC12, chalcone synthase (CHS) and flavonol synthase (FLS), while negative correlation was observed between the expression of MYB44 and the synthases; The expression of LSD1 was highly correlated with the expression of SOD and the total antioxidant capacity, while the expression of LOL1 and LOL2 exhibited a negative correlation with them; Many TFs in NAC and WRKY families may be potentially involved in the senescence process regulated by hormones and reactive oxygen species (ROS). The expression of NAC19, NAC29, and NAC53 exhibited a positive correlation with the contents of ABA and H2O2, while the expression of WRKY53, WRKY54, and WRKY70 exhibited a negative correlation with the contents of JA, SA and ABA.

    CONCLUSIONS: Our study provided a comprehensive characterization of the expression profiles of TFs during the developmental stages of L. japonica. In addition, we detected the key TFs that may play significant roles in controlling active components biosynthesis, antioxidant activity and flower senescence in L. japonica, thereby providing valuable insights into the molecular networks underlying L. japonica flower development.

    Matched MeSH terms: Acyltransferases
  10. High PHA density fed-batch cultivation strategies for 4HB-rich P(3HB-co-4HB) copolymer production by transformant Cupriavidus malaysiensis USMAA1020
    Norhafini H, Huong KH, Amirul AA
    Int J Biol Macromol, 2019 Mar 15;125:1024-1032.
    PMID: 30557643 DOI: 10.1016/j.ijbiomac.2018.12.121
    P(3HB-co-4HB) with a high 4HB monomer composition was previously successfully produced using the transformant Cupriavidus malaysiensis USMAA1020 containing an additional copy of the PHA synthase gene. In this study, high PHA density fed-batch cultivation strategies were developed for such 4HB-rich P(3HB-co-4HB). The pulse, constant and mixed feeding strategies resulted in high PHA accumulation, with a PHA content of 74-92 wt% and 4HB monomer composition of 92-99 mol%. The pulse-feed of carbon and nitrogen resulted in higher PHA concentration (30.7 g/L) than carbon alone (22.3 g/L), suggesting that a trace amount of nitrogen is essential to support cell density for PHA accumulation. Constant feeding was found to be a more feasible strategy than mixed feeding, since the latter caused a drastic fluctuation in the C/N ratio, as evidenced by higher biomass formation indicating more carbon flux towards the competitive TCA pathway. A two-times carbon and nitrogen pulse feeding was the most optimal strategy achieving 92 wt% accommodation of the total biomass, with the highest PHA concentration (46 g/L) and yield (Yp/x) of 11.5 g/g. The strategy has kept the C/N at optimal ratio during the active PHA-producing phase. This is the first report of the production of high PHA density for 4HB-rich P(3HB-co-4HB).
    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism*
  11. PHA synthase (PhaC): interpreting the functions of bioplastic-producing enzyme from a structural perspective
    Chek MF, Hiroe A, Hakoshima T, Sudesh K, Taguchi S
    Appl Microbiol Biotechnol, 2019 Feb;103(3):1131-1141.
    PMID: 30511262 DOI: 10.1007/s00253-018-9538-8
    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.
    Matched MeSH terms: Acyltransferases
  12. Modelling of polyhydroxyalkanoate synthase from Aquitalea sp. USM4 suggests a novel mechanism for polymer elongation
    Teh AH, Chiam NC, Furusawa G, Sudesh K
    Int J Biol Macromol, 2018 Nov;119:438-445.
    PMID: 30048726 DOI: 10.1016/j.ijbiomac.2018.07.147
    Polyhydroxyalkanoate (PHA) synthase, PhaC, is a key enzyme in the biosynthesis of PHA, a type of bioplastics with huge potential to replace petroleum-based plastics. While two structures have been determined, the exact mechanism remains unclear partly due to the absence of a tunnel for product passage. A model of the class I PhaC from Aquitalea sp. USM4, characterised with Km of 394 μM and kcat of 476 s-1 on 3-(R)-hydroxybutyryl-CoA, revealed a three-branched channel at the dimeric interface. Two of them are opened to the solvent and are expected to serve as the putative routes for substrate entrance and product exit, while the third is elongated in the class II PhaC1 model from Pseudomonas aeruginosa, indicating a role in accommodating the hydroxyalkanoate (HA) moiety of a HA-CoA substrate. Docking of the two tetrahedral intermediates, formed during the transfer of the growing PHA chain from the catalytic Cys to a new molecule of substrate and back to Cys, suggests a common elongation mechanism requiring the HA moiety of the ligand to rotate ~180°. Substrate specificity is determined in part by a bulky Phe/Tyr/Trp residue in the third branch in class I, which is conserved as Ala in class II to create room for longer substrates.
    Matched MeSH terms: Acyltransferases/metabolism*; Acyltransferases/chemistry*
  13. Fulltext Proof of concept in utilizing in-trans surface display system of Lactobacillus plantarum as mucosal tuberculosis vaccine via oral administration in mice
    Mustafa AD, Kalyanasundram J, Sabidi S, Song AA, Abdullah M, Abdul Rahim R, et al.
    BMC Biotechnol, 2018 10 11;18(1):63.
    PMID: 30309359 DOI: 10.1186/s12896-018-0461-y
    BACKGROUND: Tuberculosis is one of the most common and deadliest infectious diseases worldwide affecting almost a third of the world's population. Although this disease is being prevented and controlled by the Bacille Calmette Guérin (BCG) vaccine, the protective efficacy is highly variable and substandard (0-80%) in adults. Therefore, novel and effective tuberculosis vaccine that can overcome the limitations from BCG vaccine need to be developed.

    RESULTS: A novel approach of utilizing an in-trans protein surface display system of Lactobacillus plantarum carrying and displaying combination of Mycobacterium tuberculosis subunit epitope antigens (Ag85B, CFP-10, ESAT-6, Rv0475 and Rv2031c) fused with LysM anchor motif designated as ACERL was constructed, cloned and expressed in Esherichia coli Rossetta expression host. Subsequently the binding capability of ACERL to the cell wall of L. plantarum was examined via the immunofluorescence microscopy and whole cell ELISA where successful attachment and consistent stability of cell wall binding up to 4 days was determined. The immunization of the developed vaccine of L. plantarum surface displaying ACERL (Lp ACERL) via the oral route was studied in mice for its immunogenicity effects. Lp ACERL immunization was able to invoke significant immune responses that favor the Th1 type cytokine response of IFN-γ, IL-12 and IL-2 as indicated by the outcome from the cytokine profiling of spleen, lung, gastrointestinal tract (GIT), and the re-stimulation of the splenocytes from the immunized mice. Co-administration of an adjuvant consisting of Lactococcus lactis secreting mouse IL-12 (LcIL-12) with Lp ACERL was also investigated. It was shown that the addition of LcIL-12 was able to further generate significant Th1 type cytokines immune responses, similar or better than that of Lp ACERL alone which can be observed from the cytokine profiling of the immunized mice's spleen, lung and GIT.

    CONCLUSIONS: This study represents a proof of concept in the development of L. plantarum as a carrier for a non-genetically modified organism (GMO) tuberculosis vaccine, which may be the strategy in the future for tuberculosis vaccine development.

    Matched MeSH terms: Acyltransferases/administration & dosage; Acyltransferases/genetics; Acyltransferases/immunology
  14. Amphicnemis valentini sp. nov. from the Cardamom ecoregion in Cambodia and Vietnam (Odonata: Coenagrionidae)
    Kosterin OE, Kompier T
    Zootaxa, 2018 Jun 06;4429(2):281-294.
    PMID: 30313268 DOI: 10.11646/zootaxa.4429.2.4
    Amphicnemis valentini sp. nov. is described from the Ream Peninsula of Cambodia (holotype: Cambodia, Preah Sihanouk Province, Ream Peninsula, 10.52258 N 103.69556 E, RMNH) and Phú Quốc Island, Kien Giang Province of Vietnam, both in the Cardamom ecoregion. It is similar to A. gracilis Krüger, 1898, which occurs in Peninsular Malaysia and Sumatra, but differs from it by a long process on the male prothorax.
    Matched MeSH terms: Acyltransferases
  15. Fulltext Data on partial polyhydroxyalkanoate synthase genes (phaC) mined from Aaptos aaptos marine sponge-associated bacteria metagenome
    Amelia TSM, Amirul AA, Bhubalan K
    Data Brief, 2018 Feb;16:75-80.
    PMID: 29188224 DOI: 10.1016/j.dib.2017.11.011
    We report data associated with the identification of three polyhydroxyalkanoate synthase genes (phaC) isolated from the marine bacteria metagenome of Aaptos aaptos marine sponge in the waters of Bidong Island, Terengganu, Malaysia. Our data describe the extraction of bacterial metagenome from sponge tissue, measurement of purity and concentration of extracted metagenome, polymerase chain reaction (PCR)-mediated amplification using degenerate primers targeting Class I and II phaC genes, sequencing at First BASE Laboratories Sdn Bhd, and phylogenetic analysis of identified and known phaC genes. The partial nucleotide sequences were aligned, refined, compared with the Basic Local Alignment Search Tool (BLAST) databases, and released online in GenBank. The data include the identified partial putative phaC and their GenBank accession numbers, which are Rhodocista sp. phaC (MF457754), Pseudomonas sp. phaC (MF437016), and an uncultured bacterium AR5-9d_16 phaC (MF457753).
    Matched MeSH terms: Acyltransferases
  16. Fulltext In vitro and in silico studies of chalcone synthase variant 2 in Boesenbergia rotunda and its substrate specificity
    Sanmugavelan R, Teoh TC, Roslan N, Mohamed Z
    Turk J Biol, 2018;42(3):213-223.
    PMID: 30814883 DOI: 10.3906/biy-1710-107
    In this study, transformation of BrCHS var 2 into B. rotunda cell suspension culture, followed by chalcone synthase enzymatic assay and HPLC analysis was conducted to investigate whether the substrate specificity for BrCHS var 2 is either cinnamoyl-CoA or p-coumaroyl-CoA. The HPLC profile showed an increase in the amount of pinocembrin chalcone when cinnamoyl-CoA and malonyl-CoA were added but not p-coumaroyl-CoA. Molecular docking was performed to explore the binding of cinnamoyl-CoA and p-coumaroyl-CoA to BrCHS var 2 receptor and the docking results showed that cinnamoyl-CoA formed numerous hydrogen bonds and more negative docked energy than p-coumaroyl-CoA. Cinnamoyl-CoA showed good interactions with Cys 164 to initiate the subsequent formation of pinocembrin chalcone, whereas the hydroxyl group of p-coumaroyl-CoA formed an unfavorable interaction with Gln 161 that caused steric hindrance to subsequent formation of naringenin chalcone. Docked conformation analysis results also showed that malonyl-CoA formed hydrogen bonding with Cys 164, His 303, and Asn 336 residues in BrCHS var 2. The results show that cinnamoyl-CoA is the preferred substrate for BrCHS var 2.
    Matched MeSH terms: Acyltransferases
  17. Biosynthesis of P(3HB-co-3HHx) with improved molecular weights from a mixture of palm olein and fructose by Cupriavidus necator Re2058/pCB113
    Murugan P, Gan CY, Sudesh K
    Int J Biol Macromol, 2017 Sep;102:1112-1119.
    PMID: 28476592 DOI: 10.1016/j.ijbiomac.2017.05.006
    A combination of palm olein (POl) and fructose was used as carbon source for the biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by recombinant Cupriavidus necator Re2058/pCB113. Cultures grown using 5g/L PO alone as carbon source produced cell dry weight (CDW) of 5.13g/L, 67% PHA/CDW and accumulated a copolymer containing 27mol% 3HHx in shake flask cultures. When cultures were grown in 5g/L fructose alone as the carbon source they produced CDW of 2.32g/L, 11% PHA/CDW and accumulated only poly(3-hydroxybutyrate) [P(3HB)] homopolymer. When the cells were cultured in 5g/L POl in combination with 7g/L fructose, CDW of 7.41g/L and 80% PHA/CDW was obtained with 17mol% 3HHx monomer fraction. Biosynthesis was carried out using a 13L fermenter to study the accumulation of 3HHx monomer fraction in the bacterial cells at different time point. The molecular weights of P(3HB-co-3HHx) with 4-15mol% 3HHx monomer were in the range between 5.47-6.85×105Da, which were at least two fold higher than previously reported values. Interestingly, the increase in Mwof the copolymer along with the increase in 3HHx molar fraction was observed. The viscoelastic property of the copolymer further confirmed the increase in Mw.
    Matched MeSH terms: Acyltransferases/metabolism
  18. Fulltext Structure of polyhydroxyalkanoate (PHA) synthase PhaC from Chromobacterium sp. USM2, producing biodegradable plastics
    Chek MF, Kim SY, Mori T, Arsad H, Samian MR, Sudesh K, et al.
    Sci Rep, 2017 07 13;7(1):5312.
    PMID: 28706283 DOI: 10.1038/s41598-017-05509-4
    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, PhaC Cs -CAT. The structure shows that PhaC Cs -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 (PhaC Cn -CAT). Structural comparison showed PhaC Cn -CAT adopting a partially open form maintaining a narrow substrate access channel to the active site, but no product egress. PhaC Cs -CAT forms a face-to-face dimer mediated by the CAP subdomains. This arrangement of the dimer is also distinct from that of the PhaC Cn -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.
    Matched MeSH terms: Acyltransferases/metabolism; Acyltransferases/chemistry*
  19. Synthesis of high 4-hydroxybutyrate copolymer by Cupriavidus sp. transformants using one-stage cultivation and mixed precursor substrates strategy
    Syafiq IM, Huong KH, Shantini K, Vigneswari S, Aziz NA, Amirul AA, et al.
    Enzyme Microb Technol, 2017 Mar;98:1-8.
    PMID: 28110659 DOI: 10.1016/j.enzmictec.2016.11.011
    Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] copolymer is noted for its high biocompatibility, which makes it an excellent candidate for biopharmaceutical applications. The wild-type Cupriavidus sp. USMAA1020 strain is able to synthesize P(3HB-co-4HB) copolymers with different 4HB monomer compositions (up to 70mol%) in shaken flask cultures. Combinations of 4HB carbon precursors consisting of 1,6-hexanediol and γ-butyrolactone were applied for the production of P(3HB-co-4HB) with different 4HB molar fraction. A sharp increase in 4HB monomer composition was attained by introducing additional copies of PHA synthase gene (phaC), responsible for P(3HB-co-4HB) polymerization. The phaC of Cupriavidus sp. USMAA1020 and Cupriavidus sp. USMAA2-4 were cloned and heterologously introduced into host, wild-type Cupriavidus sp. USMAA1020. The gene dosage treatment resulted in the accumulation of 93mol% 4HB by the transformant strains when grown in similar conditions as the wild-type USMAA1020. The PHA synthase activities for both transformants were almost two-fold higher than the wild-type. The ability of the transformants to produce copolymers with high 4HB monomer composition was also tested in large scale production system using 5L and 30L bioreactors with a constant oxygen mass transfer rate. The 4HB monomer composition could be maintained at a range of 83-89mol%. The mechanical and thermal properties of copolymers improved with increasing 4HB monomer composition. The copolymers produced could be tailored for specific biopharmaceutical applications based on their properties.
    Matched MeSH terms: Acyltransferases/genetics; Acyltransferases/metabolism
  20. Fulltext Complete Genome Sequences of Three Cupriavidus Strains Isolated from Various Malaysian Environments
    Shafie NA, Lau NS, Ramachandran H, Amirul AA
    Genome Announc, 2017 Jan 19;5(3).
    PMID: 28104662 DOI: 10.1128/genomeA.01498-16
    Cupriavidus sp. USMAA1020, USMAA2-4, and USMAHM13 are capable of producing polyhydroxyalkanoate (PHA). This biopolymer is an alternative solution to synthetic plastics, whereby polyhydroxyalkanoate synthase is the key enzyme involved in PHA biosynthesis. Here, we report the complete genomes of three Cupriavidus sp. strains: USMAA1020, USMAA2-4, and USMAHM13.
    Matched MeSH terms: Acyltransferases
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