Proteomics analysis reveals the importance of transcriptional regulator slyA in regulation of several physiological functions in Aeromonas hydrophila

The Lysine Acetylation Modification in the Porin Aha1 of Aeromonas hydrophila Regulates the Uptake of Multidrug Antibiotics

“…After a 12% SDS-PAGE electrophoresis, the gel stained with Coomassie bright blue, for whole-cell and Kac modified protein digestion, a total of 12 mg of each sample was added to the final concentration of 10 mM DTT at 37 °C for 2.5 h. Next, we added the final concentration of 50 mM iodoacetamide to the samples and kept them in the dark for 30 min. Then, the concentration of UA buffer was diluted to 1.5 M using five times the volume of water, and trypsin (Promega) was added at a ratio of 1:50 to digest at 37 °C for 18 h. Finally, the digested peptides were desalted by SPE C18 column (Waters WAT051910) and lyophilized by CentriVap concentrator (Labconco Inc) for future mass spectrometry analysis ( 16 ).…”

Proteomics analysis reveals that CirA in Aeromonas hydrophila is involved in nutrient uptake

“…LB plates containing 0.3% (W/V) or 0.5% (W/V) agar were prepared for swimming and swarming motility assays, respectively (Li et al, 2021). Then, 2 μl of overnight‐cultured bacterial strains was spotted onto the middle of the agar plates and incubated at 30°C for 6–8 h. Swimming or swarming halo diameters were measured for three independent experiments.…”

“…Each sample was replicated across eight wells per plate as technical replicates, in addition to three independent biological replicates. Assays for extracellular protease and haemolytic activities were performed as previously described (Li et al, 2021). Briefly, 0.5-cm-diameter holes were punched at the centre of LB agar plates containing 1% skim milk or 5% sheep's blood, followed by the addition of 10 μl of overnight-cultured bacteria to the center of the hole and incubation at 30 C for 16 h. Extracellular protease (ECPase) and hemolytic activity measurements were then conducted.…”