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Repetitive Synthesis of High-Molecular-Weight Hyaluronic Acid with Immobilized Enzyme Cascades.

ABSTRACT: Industrial hyaluronic acid (HA) production comprises either fermentation with Streptococcus strains or extraction from rooster combs. The hard-to-control product quality is an obstacle to these processes. Enzymatic syntheses of HA were developed to produce high-molecular-weight HA with low dispersity. To facilitate enzyme recovery and biocatalyst re-use, here the immobilization of cascade enzymes onto magnetic beads was used for the synthesis of uridine-5'-diphosphate-α-d-N-acetyl-glucosamine (UDP-GlcNAc), UDP-glucuronic acid (UDP-GlcA), and HA. The combination of six enzymes in the UDP-sugar cascades with integrated adenosine-5'-triphosphate-regeneration reached yields between 60 and 100 % for 5 repetitive batches, proving the productivity. Immobilized HA synthase from Pasteurella multocida produced HA in repetitive batches for three days. Combining all seven immobilized enzymes in a one-pot synthesis, HA production was demonstrated for three days with a HA concentration of up to 0.37 g L^-1 , an average M_W of 2.7-3.6 MDa, and a dispersity of 1.02-1.03.

SUBMITTER: Gottschalk J

PROVIDER: S-EPMC9290584 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature

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Repetitive Synthesis of High-Molecular-Weight Hyaluronic Acid with Immobilized Enzyme Cascades.

Gottschalk Johannes J Aßmann Miriam M Kuballa Jürgen J Elling Lothar L

ChemSusChem 20210712 9

Industrial hyaluronic acid (HA) production comprises either fermentation with Streptococcus strains or extraction from rooster combs. The hard-to-control product quality is an obstacle to these processes. Enzymatic syntheses of HA were developed to produce high-molecular-weight HA with low dispersity. To facilitate enzyme recovery and biocatalyst re-use, here the immobilization of cascade enzymes onto magnetic beads was used for the synthesis of uridine-5'-diphosphate-α-d-N-acetyl-glucosamine (U ...[more]

PMID: 34143936

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Project description:BackgroundThe rapid decline in ovarian function associated with menopause promotes osteoclast differentiation and increases bone resorption, disrupting of bone homeostasis and increasing the risk of osteoporosis. Hyaluronic acid (HA) is a polysaccharide ubiquitously present in the connective tissues. Recent reports indicate that high-molecular-weight HA (HMW-HA) promotes osteoblast proliferation, enhances alkaline phosphatase activity and mineral deposition, and promotes the expression of bone differentiation markers, such as Runx2 and osteocalcin. HMW-HA also inhibits the expression of the receptor activator of nuclear factor kappa-B ligand (RANKL) in osteoblasts. These results suggest that HMW-HA may be an effective therapeutic agent against postmenopausal osteoporosis. Therefore, this study aimed to examine whether HMW-HA alleviates ovariectomy (OVX)-induced bone loss in mice.MethodsEight-week-old female C57BL/6 J mice were randomly divided into the following five groups: Group 1: Sham/saline, Group 2: OVX/saline, Group 3: OVX/HMW-HA [15 mg/kg]; Group 4: OVX/HMW-HA [30 mg/kg]; and Group 5: OVX/HMW-HA [60 mg/kg]. Mice were administered HMW-HA or saline subcutaneously starting from 1 week after OVX and changes in bone mass were analyzed at 5 weeks using three-dimensional micro-computed tomography (3D-μCT). In addition, changes in osteoclast parameters were analyzed histologically.ResultsThe reduction in trabecular bone volume and trabecular number was significantly ameliorated in the OVX/HMW-HA group compared with that observed in the OVX/saline group, along with a significant inhibition of the increase in trabecular spacing. In addition, the OVX/HMW-HA group exhibited a significant reduction in osteoclast surface area and number compared with the OVX/saline group, with no significant differences compared with the sham group. In vitro experiments revealed that depletion of HMW-HA from the culture medium by hyaluronidase treatment increased RANKL expression in the bone marrow stromal cell line ST2. These data suggest that HMW-HA alleviates OVX-induced bone loss by downregulating osteoclast formation and/or activity in mice.ConclusionHMW-HA is a potential novel therapeutic agent for osteoporosis.

Dataset Information

Repetitive Synthesis of High-Molecular-Weight Hyaluronic Acid with Immobilized Enzyme Cascades.

Publications

Repetitive Synthesis of High-Molecular-Weight Hyaluronic Acid with Immobilized Enzyme Cascades.

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