Engineering Halomonas bluephagenesis TD01 as a Robust Chassis for the Sustainable Production of Hyaluronic Acid

This study evaluates the development of Halomonas bluephagenesis TD01 as a novel, sustainable microbial platform for the production of hyaluronic acid (HA). Three distinct hyaluronan synthase genes (sezHasA and spHasA "Class I" from the Streptococcal group and pmHasA "class II") were heterologously expressed and compared, with the Class II synthase from Pasteurella multocida (pmHasA) emerging as the superior variant in rich media 60-LBG, achieving significantly higher titers of 0.88 g/L and molecular weight (Mw) of 1.15 MDa (Mega Daltons). Using a combination of Plackett–Burman design and Response Surface Methodology (RSM), the fermentation process was optimized, identifying initial pH, nitrogen source, and NaCl concentration as critical factors. These optimizations led to a maximum HA yield from 0.88 to 2.38 g/L (265% improvement) and Mw from 1.15 to 9.67 MDa. Furthermore, the study demonstrates precise tuning of HA molecular weight, ranging from 2.04 MDa to 9.67 MDa in a modified medium (40LBG-Y), by modulating L-arabinose induction levels. The structural integrity of the purified HA was confirmed via ESI-MS and 1H-NMR. These findings establish H. bluephagenesis TD01 as a robust Next-Generation Industrial Biotechnology (NGIB) chassis for the scalable and customizable production of HA with a minimal cost and high-molecular-weight HA for medical applications.