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Linalool and nerolidol are terpene alcohols that occur naturally in many aromatic plants and are commonly used in food and cosmetic industries as flavors and fragrances. In plants, linalool and nerolidol are biosynthesized as a result of respective linalool synthase and nerolidol synthase, or a single linalool/nerolidol synthase. In our previous work, we have isolated a linalool/nerolidol synthase (designated as PamTps1) from a local herbal plant, Plectranthus amboinicus, and successfully demonstrated the production of linalool and nerolidol in an Escherichia coli system. In this work, the biochemical properties of PamTps1 were analyzed, and its 3D homology model with the docking positions of its substrates, geranyl pyrophosphate (C10) and farnesyl pyrophosphate (C15) in the active site were constructed. PamTps1 exhibited the highest enzymatic activity at an optimal pH and temperature of 6.5 and 30 °C, respectively, and in the presence of 20 mM magnesium as a cofactor. The Michaelis–Menten constant (Km) and catalytic efficiency (kcat/Km) values of 16.72 ± 1.32 µM and 9.57 × 10–3 µM−1 s−1, respectively, showed that PamTps1 had a higher binding affinity and specificity for GPP instead of FPP as expected for a monoterpene synthase. The PamTps1 exhibits feature of a class I terpene synthase fold that made up of α-helices architecture with N-terminal domain and catalytic C-terminal domain. Nine aromatic residues (W268, Y272, Y299, F371, Y378, Y379, F447, Y517 and Y523) outlined the hydrophobic walls of the active site cavity, whilst residues from the RRx8W motif, RxR motif, H-α1 and J-K loops formed the active site lid that shielded the highly reactive carbocationic intermediates from the solvents. The dual substrates use by PamTps1 was hypothesized to be possible due to the architecture and residues lining the catalytic site that can accommodate larger substrate (FPP) as demonstrated by the protein modelling and docking analysis. This model serves as a first glimpse into the structural insights of the PamTps1 catalytic active site as a multi-substrate linalool/nerolidol synthase. [ABSTRACT FROM AUTHOR]

Lignosulfonate (LS) is a commonly used to promote plant growth. However, the underlying growth promoting responses of LS in plant remain unknown. Therefore, this study was undertaken to elucidate the underlying growth promoting mechanisms of LS, specifically calcium lignosulfonate (CaLS). Addition of 100 mg/L CaLS in phytohormone-free media enhanced recalcitrant indica rice cv. MR219 callus proliferation rate and adventitious root formation. Both, auxin related genes (OsNIT1 , OsTAA1 and OsYUC1) and tryptophan biosynthesis proteins were upregulated in CaLS-treated calli which corroborated with increased of endogenous auxin content. Moreover, increment of OsWOX11 gene on CaLS-treated calli implying that the raised of endogenous auxin was utilized as a cue to enhance adventitious root development. Besides, CaLS-treated calli showed higher nutrient ions content with major increment in calcium and potassium ions. Consistently, increased of potassium protein kinases genes (OsAKT1 , OsHAK5 , OsCBL , OsCIPK23 and OsCamk1) were also recorded. In CaLS treated calli, the significant increase of calcium ion was observed starting from week one while potassium ion only recorded significant increase on week two onwards, suggesting that increment of potassium ion might be dependent on the calcium ion content in the plant cell. Additionally, reduced callus blackening was also coherent with downregulation of ROS scavenging protein and reduced H 2 O 2 content in CaLS-treated calli suggesting the role of CaLS in mediating cellular homeostasis via prevention of oxidative burst in the cell. Taken together, CaLS successfully improved MR219 callus proliferation and root formation by increasing endogenous auxin synthesis, enhancing nutrients uptake and regulating cellular homeostasis. • CaLS stimulates root formation in rice callus via regulation of auxin biosynthesis. • Enhancement of nutrient uptake was observed in CaLS-treated rice calli. • CaLS improves cellular homeostasis level and enhances callus proliferation rate. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae (KP) remains the most prevalent nosocomial pathogen and carries the carbapenemase (KPC) gene which confers resistance towards carbapenem. Thus, it is necessary to discover novel antimicrobials to address the issue of antimicrobial resistance in such pathogens. Natural products such as essential oils are a promising source due to their complex composition. Essential oils have been shown to be effective against pathogens, but the overall mechanisms have yet to be fully explained. Understanding the molecular mechanisms of essential oil towards KPC-KP cells would provide a deeper understanding of their potential use in clinical settings. Therefore, we aimed to investigate the mode of action of essential oil against KPC-KP cells from a proteomic perspective by comparing the overall proteome profile of KPC-KP cells treated with cinnamon bark (Cinnamomum verum J. Presl) essential oil (CBO) at their sub-inhibitory concentration of 0.08% (v/v). A total of 384 proteins were successfully identified from the non-treated cells, whereas only 242 proteins were identified from the CBO-treated cells. Proteins were then categorized based on their biological processes, cellular components and molecular function prior to pathway analysis. Pathway analysis showed that CBO induced oxidative stress in the KPC-KP cells as indicated by the abundance of oxidative stress regulator proteins such as glycyl radical cofactor, catalase peroxidase and DNA mismatch repair protein. Oxidative stress is likely to oxidize and disrupt the bacterial membrane as shown by the loss of major membrane proteins. Several genes selected for qRT-PCR analysis validated the proteomic profile and were congruent with the proteomic abundance profiles. In conclusion, KPC-KP cells exposed to CBO undergo oxidative stress that eventually disrupts the bacterial membrane possibly via interaction with the phospholipid bilayer. Interestingly, several pathways involved in the bacterial membrane repair system were also affected by oxidative stress, contributing to the loss of cells viability. [ABSTRACT FROM AUTHOR]