Low molecular weight heparosan is an un-sulfated polysaccharide primarily used as a precursor for heparin synthesis that has recently been used in drug delivery applications. Heparosan synthesis from recombinant bacterial systems provides a safer alternative to naturally producing pathogenic bacterial systems. In this study, we engineered a functional heparosan synthesis pathway in Bacillus megaterium by the expression of E. coli K5 kfiC and kfiA glycosyltransferase genes. Upregulation of individual UDP-sugar precursor pathway genes enhanced the heparosan production, indicating that UDP-precursor sugar concentrations were limiting the biosynthesis. The engineered B. megaterium yielded a maximum heparosan concentration of 394 mg/L in batch bioreactor. The heparosan titer was further increased to 1.32 g/L in fed-batch fermentation. Nuclear magnetic resonance analysis revealed that the chemical structure of B. megaterium derived heparosan was identical to E. coli K5 heparosan. The heparosan molecular weight varied from 31 to 60 kDa, indicating its potential as a precursor for chemoenzymatic heparin biosynthesis. This study provides an efficient process to produce heparosan from non-pathogenic B. megaterium.
glycosyltransferases, Heparosan, Bacillus megaterium
NCBI PubMed ID: 32445821Publication DOI: 10.1016/j.ijbiomac.2020.05.159Journal NLM ID: 7909578Publisher: Butterworth-Heinemann
Correspondence: senthilkumar@iitg.ac.in
Institutions: Bioprocess Analytical Technology Laboratory, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Assam, India, Molecular Endocrinology Laboratory, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Assam, India
Methods: 13C NMR, 1H NMR, genetic methods, HPSEC, fermentation
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