Recent advancements in biotechnology have unlocked new possibilities for the production of hyaluronic acid (HA), a widely used compound with applications ranging from cosmetics to pharmaceuticals. Researchers have engineered variants of hyaluronic acid synthase (HAS) to successfully produce HA with controllable molecular weights, overcoming historical challenges associated with its high-yield synthesis.
Hyaluronic acid is notable for its unique structural properties; it is composed of repeating disaccharide units of N-acetylglucosamine and glucuronic acid. Its varying molecular weights (MWs) significantly influence its physicochemical properties and biological functions. For example, high MW HA is utilized for its viscoelastic properties, making it ideal for osteoarthritis treatments, whereas low MW HA has applications ranging from drug delivery to influencing inflammation.
A recent study focused on the heterologous production of HA in the bacterium Corynebacterium glutamicum, which has garnered attention due to its Generally Recognized as Safe (GRAS) status and flexible metabolic capabilities. Traditionally, controlling HA biosynthesis has proven difficult due to the metabolic imbalances between the biosynthetic pathways and cellular growth rates. Addressing this, the research team aimed to understand the structure-function relationship of Class I HA synthases and manipulate them for enhanced production.
The authors implemented several key strategies to optimize HA production. They systematically characterized Class I HAS genes from Streptococcus equi subsp. zooepidemicus, pinpointing regions of the enzymes important for HA polymerization and secretion. After constructing HAS mutants, the researchers achieved complete HA secretion from engineered C. glutamicum cells, significantly extending the molecular weight range of produced HA from 300 kDa to 1400 kDa.
One of the groundbreaking aspects of this research centered around dynamic regulation of the enzyme UDP-glucose 6-dehydrogenase, which is pivotal for modulating the metabolic pathways involved in HA production. By fine-tuning its activity and employing adaptive laboratory evolution strategies, the scientists ensured the normal growth of C. glutamicum cells, even under conditions typically detrimental to cellular health due to high viscosity from accumulated HA.
Final titers and productivities within this engineered system reached impressive levels: concentrations of 45 g/L for high MW HA (500 kDa) and 105 g/L for low MW HA (10 kDa) were achieved, representing significant improvements over previous attempts. "Our findings advance our knowledge of HAS function and the interplay between cell metabolism and morphology, providing insights for optimizing microbial factories for HA production," noted the authors.
The study revealed not only the structural functions of HAS but also highlighted the surprising impact of cell morphology. During the engineering process, the transformation of C. glutamicum cell morphology was observed, shifting from standard rod shapes to more irregular forms as HA accumulated. This morphological change correlated with modifications to the bacterial membrane composition, underscoring the role of lipid interactions with membrane proteins.
Performance testing of the engineered strains, particularly C. glutamicum CGHA-125, showcased the potential for industrial application. This strain adapted effectively under high viscus conditions, achieving remarkable yields and paving the way for commercial HA production systems. The adaptive laboratory evolution strategy applied allowed researchers to continuously select for strains with improved HA synthesis capabilities, enhancing metabolic capacity.
Concluding, this innovative research heralds significant shifts for the biosynthesis of complex polysaccharides. With improved insights and strategies for HA production, the study sets the stage for future advancements within microbial manufacturing, promising scalable and efficient solutions for various biomedical applications, said the authors. This progress reflects the continuous evolution of methodologies to equip synthetic biology with tools needed for high-caliber production demands.