Heavy Heparin: A Stable Isotope‐Enriched, Chemoenzymatically‐Synthesized, Poly‐Component Drug

Cress, Brady F.; Bhaskar, Ujjwal; Vaidyanathan, Deepika; Williams, Arthur Robin; Cai, Chao; Liu, Xinyue; Fu, Li; M‐Chari, Vandhana; Zhang, Fuming; Mousa, Shaker A.; Dordick, Jonathan S.; Koffas, Mattheos; Linhardt, Robert J.

doi:10.1002/anie.201900768

Cited by 41 publications

(24 citation statements)

References 35 publications

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“…This one-step, sustainable process offers a promising alternative to animal extraction in GAG production. This approach also offers the potential to prepare unnatural GAG derivatives and stable isotope-labeled GAGs 45 . In improving the precursors required for CS synthesis, we also demonstrate an increased sulfation level of the intracellular product to ~96%, comparable to commercially available CS-A.…”

Section: Discussionmentioning

confidence: 99%

Complete biosynthesis of a sulfated chondroitin in Escherichia coli

et al. 2021

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Sulfated glycosaminoglycans (GAGs) are a class of important biologics that are currently manufactured by extraction from animal tissues. Although such methods are unsustainable and prone to contamination, animal-free production methods have not emerged as competitive alternatives due to complexities in scale-up, requirement for multiple stages and cost of co-factors and purification. Here, we demonstrate the development of single microbial cell factories capable of complete, one-step biosynthesis of chondroitin sulfate (CS), a type of GAG. We engineer E. coli to produce all three required components for CS production–chondroitin, sulfate donor and sulfotransferase. In this way, we achieve intracellular CS production of ~27 μg/g dry-cell-weight with about 96% of the disaccharides sulfated. We further explore four different factors that can affect the sulfation levels of this microbial product. Overall, this is a demonstration of simple, one-step microbial production of a sulfated GAG and marks an important step in the animal-free production of these molecules.

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Section: Discussionmentioning

confidence: 99%

Complete biosynthesis of a sulfated chondroitin in Escherichia coli

et al. 2021

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“…Non-anticoagulant low molecular weight HP (NACH) was synthesized from dalteparin, a nitrous acid depolymerization product of porcine intestinal HP, followed by periodate oxidation 25 . Non-anticoagulant heparin TriS (NS2S6S) was synthesized from N -sulfo heparosan with subsequent modification with C5-epimerase and 2- O - and 6- O -sulfotransferases (2OST and 6OST1/6OST3) 26 . Heparin oligosaccharides included tetrasaccharide, hexasaccharide, octasaccharide, decasaccharide, dodecasaccharide, tetradecasaccharide, hexadecasaccharide and octadecasaccharide and were prepared from porcine intestinal heparin via controlled partial heparin lyase 1 treatment followed by size fractionation.…”

Section: Methodsmentioning

confidence: 99%

Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives

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Zhang

et al. 2020

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Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic or prophylactic. Like other betacoronaviruses, attachment and entry of SARS-CoV-2 is mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in anti-viral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH and 6-O-desulfated enoxaparin with an IC50 of 599 ng/L, 108 µg/L, 177 ng/L, and 586 µg/L respectively. The low serum bioavailability of intranasally administered UFH, along with data suggesting that the nasal epithelium is a portal for initial infection and transmission, suggest that intranasal administration of UFH may be an effective and safe prophylactic treatment.

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“…For each set of competition experiments, a control experiment (S-protein without polysaccharide) was performed to ensure the surface was fully regenerated. Among the tested polysaccharides, RPI-27 and RPI-28, complex sulfated polysaccharides (fucoidans) extracted from the seaweed Saccharina japonica 6 , chemo-enzymatically synthesized trisulfated (TriS) heparin 7 , and unfractionated USP-heparin itself were able to compete with heparin for S-protein binding. We selected these compounds along with a non-anticoagulant low molecular weight heparin (NACH) 8 for further study (Fig.…”

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confidence: 99%