Rhamnose

The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall teichoic acids (WTA). The presence and chemical structure of many rhamnose-containing cell wall polysaccharides (RhaCWP) has sometimes been known for decades. In contrast to WTA, insight into the biosynthesis and functional role of RhaCWP has been lacking. Recent studies in human streptococcal and enterococcal pathogens have highlighted critical roles for these complex polysaccharides in bacterial cell wall architecture and pathogenesis. In this review, we provide an overview of the RhaCWP with regards to their biosynthesis, genetics and biological function in species most relevant to human health. We also briefly discuss how increased knowledge in this field can provide interesting leads for new therapeutic compounds and improve biotechnological applications.

BackgroundA capable expression vector is mainly characterized by its production efficiency, stability and induction response. These features can be influenced by a variation of modifications and versatile genetic modules.ResultsWe examined miscellaneous variations of a rhaPBAD expression vector. The introduction of a stem loop into the translation initiation region of the rhaPBAD promoter resulted in the most significant improvement of eGFP expression. Starting from this plasmid, we constructed a set of expression vectors bearing different genetic modules like rop, ccdAB, cer and combinations thereof, and tested the efficiency of expression and plasmid stability. The plasmid pWA21, containing the stem loop, one cer site and rop, attained high expression levels accompanied by a good stability, and on that score seems to be a well-balanced choice.ConclusionWe report the generation of variations of the rhaPBAD expression vector and characterization hereof. The genetic modules showed a complex interplay, therefore two positive effects combined sometimes resulted in a disadvantage.

Listeria monocytogenes is an opportunistic Gram-positive bacterial pathogen responsible for listeriosis, a human foodborne disease. Its cell wall is densely decorated with wall teichoic acids (WTAs), a class of anionic glycopolymers that play key roles in bacterial physiology, including protection against the activity of antimicrobial peptides (AMPs). In other Gram-positive pathogens, WTA modification by amine-containing groups such as D-alanine was largely correlated with resistance to AMPs. However, in L. monocytogenes, where WTA modification is achieved solely via glycosylation, WTA-associated mechanisms of AMP resistance were unknown. Here, we show that the L-rhamnosylation of L. monocytogenes WTAs relies not only on the rmlACBD locus, which encodes the biosynthetic pathway for L-rhamnose, but also on rmlT encoding a putative rhamnosyltransferase. We demonstrate that this WTA tailoring mechanism promotes resistance to AMPs, unveiling a novel link between WTA glycosylation and bacterial resistance to host defense peptides. Using in vitro binding assays, fluorescence-based techniques and electron microscopy, we show that the presence of L-rhamnosylated WTAs at the surface of L. monocytogenes delays the crossing of the cell wall by AMPs and postpones their contact with the listerial membrane. We propose that WTA L-rhamnosylation promotes L. monocytogenes survival by decreasing the cell wall permeability to AMPs, thus hindering their access and detrimental interaction with the plasma membrane. Strikingly, we reveal a key contribution of WTA L-rhamnosylation for L. monocytogenes virulence in a mouse model of infection.

Seaweed represents a potential sustainable carbon source for industrial biotechnology, yet the workhorse bacterium Corynebacterium glutamicum cannot naturally metabolize the deoxy sugars L-rhamnose and L-fucose, abundant in green and brown seaweed, respectively. Expanding its substrate range is crucial for sustainable bioprocessing, by enabling utilization of the available biomass. In this study, we engineered C. glutamicum to utilize L-rhamnose and L-fucose by introducing the Escherichia coli operons rhaBADM and fucIKUA, enabling growth on these sugars as sole carbon sources. To enhance growth, we evaluated various transport systems and identified the non-native fucose permease (FucP) as the most efficient for L-rhamnose uptake, and the native myo-inositol transporter 2 (IolT2) as optimal for L-fucose uptake. During L-rhamnose and L-fucose utilization, L-lactaldehyde is formed as a byproduct. We demonstrate that the native acetaldehyde dehydrogenase encoded by ald also exhibits lactaldehyde dehydrogenase activity, and that its overexpression enhances L-lactaldehyde utilization. Finally, cultivation on green and brown seaweed hydrolysates enabled the engineered strains to achieve increased biomass formation through consumption of the targeted deoxy sugars. This study expands the substrate spectrum of C. glutamicum through pathway engineering, transport optimization, and functional identification of a native lactaldehyde dehydrogenase. Growth and L-lysine production on seaweed hydrolysates highlights the potential of this approach for sustainable marine biomass valorization and bioproduction of value-added compounds.