RESUMO
Mining of plant-derived antimicrobials is the major focus at current to counter antibiotic resistance.This study was conducted to characterize the antimicrobial activity and mode of action of linalyl anthranilate(LNA)against carbapenemase-producing Klebsiella pneumoniae(KPC-KP).LNA alone exhibited bacteri-cidal activity at 2.5％(V/V),and in combination with meropenem(MPM)at 1.25％(V/V).Comparative proteomic analysis showed a significant reduction in the number of cytoplasmic and membrane proteins,indicating membrane damage in LNA-treated KPC-KP cells.Up-regulation of oxidative stress regulator proteins and down-regulation of oxidative stress-sensitive proteins indicated oxidative stress.Zeta po-tential measurement and outer membrane permeability assay revealed that LNA increases both bacterial surface charge and membrane permeability.Ethidium bromide influx/efflux assay showed increased uptake of ethidium bromide in LNA-treated cells,inferring membrane damage.Furthermore,intracel-lular leakage of nucleic acid and proteins was detected upon LNA treatment.Scanning and transmission electron microscopies again revealed the breakage of bacterial membrane and loss of intracellular ma-terials.LNA was found to induce oxidative stress by generating reactive oxygen species(ROS)that initiate lipid peroxidation and damage the bacterial membrane.In conclusion,LNA generates ROS,initiates lipid peroxidation,and damages the bacterial membrane,resulting in intracellular leakage and eventually killing the KPC-KP cells.
RESUMO
Abstract Sclareol is an important intermediate for ambroxide synthesis industries. Hyphozyma roseonigra ATCC 20624 was the only reported strain capable of degrading sclareol to the main product of sclareol glycol, which is the precursor of ambroxide. To date, knowledge is lacking about the effects of sclareol on cells and the proteins involved in sclareol metabolism. Comparative proteomic analyses were conducted on the strain H. roseonigra ATCC 20624 by using sclareol or glucose as the sole carbon source. A total of 79 up-regulated protein spots with a >2.0-fold difference in abundance on 2-D gels under sclareol stress conditions were collected for further identification. Seventy spots were successfully identified and finally integrated into 30 proteins. The up-regulated proteins under sclareol stress are involved in carbon metabolism; and nitrogen metabolism; and replication, transcription, and translation processes. Eighteen up-regulated spots were identified as aldehyde dehydrogenases, which indicating that aldehyde dehydrogenases might play an important role in sclareol metabolism. Overall, this study may lay the fundamentals for further cell engineering to improve sclareol glycol production.
RESUMO
Abstract Sclareol is an important intermediate for ambroxide synthesis industries. Hyphozyma roseonigra ATCC 20624 was the only reported strain capable of degrading sclareol to the main product of sclareol glycol, which is the precursor of ambroxide. To date, knowledge is lacking about the effects of sclareol on cells and the proteins involved in sclareol metabolism. Comparative proteomic analyses were conducted on the strain H. roseonigra ATCC 20624 by using sclareol or glucose as the sole carbon source. A total of 79 up-regulated protein spots with a >2.0-fold difference in abundance on 2-D gels under sclareol stress conditions were collected for further identification. Seventy spots were successfully identified and finally integrated into 30 proteins. The up-regulated proteins under sclareol stress are involved in carbon metabolism; and nitrogen metabolism; and replication, transcription, and translation processes. Eighteen up-regulated spots were identified as aldehyde dehydrogenases, which indicating that aldehyde dehydrogenases might play an important role in sclareol metabolism. Overall, this study may lay the fundamentals for further cell engineering to improve sclareol glycol production.