## 3 Results
### Proteome Analysis after Growth on IMOS  
In order to investigate the metabolic effect of IMOS on the fitness of _B. subtilis_ CU1, we assessed a 2D-PAGE proteomic analysis of the strain after 12 h of growth on glucose or IMOS as unique substrates (\(n=3\) per group). A total of 69 spots differentially expressed between the two groups were selected with a fold-change greater than 1.3 and a \(p\)-value \(\leq 0.5\) (Student's \(t\)-test on the logged ratio), then subjected to identification by Nano-LC MALDI-TOF/TOF (Figure S3). Among them, 28 proteins were identified and are presented in Table 4. A total of 13 proteins were overexpressed and 15 under expressed in IMOS conditions. Several proteins involved in protein synthesis (elongation factor G, elongation factor Tu and chaperonin GroEL) as well as purine synthesis (glycine hydroxymethyltransferase) were identified among the under-expressed proteins. This observation could be explained by the slightly reduced growth of the bacteria at 12 h on the IMOS substrate, with a possibly reduced cellular proliferation. Several proteins involved in the metabolism and transport of carbohydrates were identified, including maltose-6-phosphate glucosidase, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, probable 6-phospho-\(\beta\)-glucosidase and fructose bisphosphate aldolase. Maltose-6-phosphate glucosidase (GlyA or MalA), which is overexpressed in the presence of IMOS, is an enzyme capable of hydrolyzing maltose-6-phosphate into glucose and glucose-6-phosphate, suggesting the upstream transformation of IMOS into maltose-6-phosphate and isomaltose-6-phosphate. Glyceraldehyde-3-phosphate dehydrogenase (GapA or GAPDH) is downregulated in the presence of IMOS. It is a key kinase of glycolysis, which enables the reversibleconversion of glyceraldehyde-3-phosphate to glyceraldehyde-1,3-bisphosphate [26] for the production of pyruvate. The reaction catalyzed by this enzyme is also responsible for the production of reducing force (NADH). The identification of a 6-phospho-\(\beta\)-glucosidase for few spots is difficult to explain. Indeed, no \(\beta\)-glycosidic bond was observed by mass spectrometry in the structure of IMOS, and the use of this enzyme for the assimilation of the prebiotic substrate has not been demonstrated. In contrast, two other glycolysis enzymes, a phosphoglycerate kinase (Pgk) and a probable fructose bisphosphate aldolase (FbaA), were under expressed in the presence of IMOS. FbaA catalyzes a reversible aldolization reaction of glyceraldehyde-3-phosphate to fructose 1,6-bisphosphate. We also observed the repression of LDH expression in the presence of IMOS, which explains the decrease in lactic acid production reported in this study (Figure 2E). UDP-glucose 4-epimerase (GalE) is overexpressed in the presence of IMOS. It is a galactose metabolic enzyme that catalyzes the reversible conversion of UDP-galactose to UDP-glucose, coupled with the reduction in NAD. UDP-glucose and UDP-galactose are nucleotides required for the biosynthesis of exopolysaccharides (EPSs) [27]. In addition, in _B. subtilis_, GalE catalyzes the interconversion of UDP-GlcNAc to UDP-GalNac [28]. This result suggests that IMOS could modulate the synthesis of EPS by _B. subtilis_ CU1, as observed on the agar plate. The expression of isocitrate dehydrogenase (icd), an enzyme involved in the Krebs cycle, is reduced in the presence of IMOSs. In bacteria, this enzyme is repressed by an isocitrate dehydrogenase kinase that inactivates the enzyme by phosphorylation when there is sufficient ATP in the cell. It seems that a second regulatory site, this time at the level of transcription or translation, can be set up by the bacteria to control ATP levels in the cell. This observation suggests that the energy yield with IMOS is greater than with glucose as the sole carbon substrate. Succinate dehydrogenase (SdhA) and malate dehydrogenase (Mdh) are two enzymes involved in the Krebs cycle.