Hyaluronic acid production and characterization by novel Bacillus subtilis harboring truncated Hyaluronan Synthase

Abstract

Hyaluronic Acid (HA) is a natural biopolymer that has important physiological and industrial applications due to its viscoelastic and hydrophilic characteristics. The responsible enzyme for HA production is Hyaluronan synthase (HAS). Although in vitro structure–function of intact HAS enzyme has been partly identified, there is no data on in vivo function of truncated HAS forms. In the current study, novel recombinant Bacillus subtilis strains harboring full length (RBSFA) and truncated forms of SeHAS (RBSTr4 and RBSTr3) were developed and HA production was studied in terms of titer, production rate and molecular weight (Mw). The maximum HA titer for RBSFA, RBSTr4 and RBSTr3 was 602 ± 16.6, 503 ± 19.4 and 728 ± 22.9 mg/L, respectively. Also, the HA production rate was 20.02, 15.90 and 24.42 mg/L.h−1, respectively. The findings revealed that RBSTr3 produced 121% and 137% more HA rather than RBSFA and RBSTr4, respectively. More interestingly, the HA Mw was about 60 kDa for all strains which is much smaller than those obtained in prior studies.

The effect of manipulating glucuronic acid biosynthetic pathway in Bacillus subtilis strain on hyaluronic acid production

“…The cultured cells were removed by centrifugation and the supernatant was subjected to purification step. HA purification was performed in five steps as described previously with minor modifications (Amjad Zanjani et al 2022 ). (1) Precipitation of protein impurities was done by the addition of trichloroacetic acid (TCA, 100% w/v) and incubation at 4ºC for 1.5 h followed by centrifugation at 5000 g for 20 min at 4ºC.…”