The role of protein modifications in senescence of freeze-dried Acetobacter senegalensis during storage

Abstract

BackgroundLoss of viability is one of the most important problems during starter culture production. Previous research has mostly focused on the production process of bacterial starters, but there are few studies about cellular protein deterioration causing cell defectiveness during storage. In the present study, we investigated the influence of storage temperature (−21, 4, 35°C) on the cellular protein modifications which may contribute to the senescence of freeze-dried Acetobacter senegalensis.ResultsHeterogeneous populations composed of culturable cells, viable but non-culturable cells (VBNC) and dead cells were generated when freeze-dried cells were kept at −21 and 4°C for 12 months whereas higher storage temperature (35°C) mainly caused death of the cells. The analysis of stored cell proteome by 2D-DiGE demonstrated a modified pattern of protein profile for cell kept at 4 and 35°C due to the formation of protein spot trains and shift of Isoelectric point (pI). Quantification of carbonylated protein by ELISA showed that the cells stored at 4 and 35°C had higher carbonylated protein contents than fresh cells. 2D-DiGE followed by Western blotting also confirmed the carbonylation of cellular proteins involved in translation process and energy generation. The auto-fluorescent feature of cells kept at 35°C increased significantly which may be an indication of protein glycation during storage. In addition, the percentage of cellular unsaturated fatty acid and the solubility of cellular proteins decreased upon storage of cells at higher temperature suggesting that peroxidation of fatty acids and possibly protein lipidation and oxidation occurred.ConclusionsHigh storage temperature induces some deteriorative reactions such as protein oxidation, lipidation and glycation which may cause further protein modifications like pI-shift, and protein insolubility. These modifications can partly account for the changes in cell viability. It can also be deduced that even moderate carbonylation of some critical cellular proteins (like ribosomal proteins) may lead to VBNC formation or death of freeze-dried bacteria. Moreover, it seems that other mechanisms of biomolecule deterioration preceding protein carbonylation lead to VBNC formation under very low storage temperature.

Simultaneous production of acetic and gluconic acids by a thermotolerant Acetobacter strain during acetous fermentation in a bioreactor

“…The compositions of liquid culture media and fermentation broth were analysed by HPLC (Agilent 1110 series; Agilent Technologies, CA, USA) using the method described by Shafiei et al (16,22).…”

Effect of Sequential Acclimation to Various Carbon Sources on the Proteome of Acetobacter senegalensis LMG 23690T and Its Tolerance to Downstream Process Stresses

“…Triplicate 2D-DiGE electrophoresis were carried out on acclimated cells according to the procedures previously described (Shafiei et al, 2014). Cells acclimated under the three different conditions (see section “Acclimation in Bioreactor”) were compared two by two.…”

“…Protein spots which were different in abundance (±1.5-fold standardized abundance, p < 0.05) were picked and collected. They were then identified by mass spectrometry as described by Shafiei et al (2014).…”

“…It is able to oxidize ethanol at 25–42°C (Ndoye et al, 2007a; Shafiei et al, 2013a). This strain was successfully used for production of vinegar and vinegar starter in pilot plant scale acetifier (Ndoye et al, 2007b; Shafiei et al, 2014). Interestingly, Acetobacter senegalensis LMG 23690 T produces acetic acid and gluconic acid simultaneously at high temperature (38°C) when grown on a mixture of ethanol and glucose (Mounir et al, 2016).…”

Modified semi‐continuous fermentation for resuscitating nongrowing cells during high‐temperature gluconic acid production by Acetobacter senegalensis

“…In the present study , Acetobacter senegalensis is used as a thermo‐tolerant acetic acid bacterium isolated from mango fruit (Ndoye et al ). It was used for vinegar production and vinegar starter production (Ndoye et al ; Shafiei et al ; Shafiei et al ; Shafiei et al ). In 2017, Shafiei and coworkers evaluated sodium gluconate production by A. senegalensis in the batch culture at 38°C; however, cells were not able neither to consume glucose nor to produce gluconic acid at the end of stationary phase (Shafiei et al ).…”