Donor substrate promiscuity of the N-acetylglucosaminyltransferase activities of Pasteurella multocida heparosan synthase 2 (PmHS2) and Escherichia coli K5 KfiA

Yanhong Li, Hai Yu, Vireak Thon, Yi Chen, Musleh M. Muthana, Jingyao Qu, Liana Hie, Xi Chen

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

The biological activities of heparan sulfate (HS) and heparin (HP) are closely related to their molecular structures. Both Pasteurella multocida heparosan synthase 2 (PmHS2) and Escherichia coli K5 KfiA have been used for enzymatic and chemoenzymatic synthesis of HS and HP oligosaccharides and their derivatives. We show here that cloning using the pET15b vector and expressing PmHS2 as an N-His6-tagged fusion protein improve its expression level in E coli. Investigation of the donor substrate specificity of the N-acetylglucosaminyltransferase activities of P. multocida heparosan synthase 2 (PmHS2) and E coli K5 KfiA indicates the substrate promiscuities of PmHS2 and KfiA. Overall, both PmHS2 and KfiA can use uridine 5'-diphosphate-N-acetylglucosamine (UDP-GlcNAc) and some of its C2'- and C6'-derivatives as donor substrates for their α1-4-GlcNAcT activities. Nevertheless, PmHS2 has a broader tolerance towards substrate modifications. Other than the UDP-sugars that can be used by KfiA, additional C6'-derivatives of UDP-GlcNAc, UDP-glucose, and UDP-N-acetylgalactosamine (UDP-GalNAc) are tolerable substrates for the α1-4-GlcNAcT activity of PmHS2. The substrate promiscuities of PmHS2 and KfiA will allow efficient chemoenzymatic synthesis of diverse HS and HP oligosaccharide derivatives which may have improved or altered activities compared to their natural counterparts.

Original languageEnglish (US)
Pages (from-to)1127-1134
Number of pages8
JournalApplied Microbiology and Biotechnology
Volume98
Issue number3
DOIs
StatePublished - 2014

Keywords

  • Escherichia coli K5 KfiA
  • GlcNAc transferase
  • Heparosan synthase
  • Pasteurella multocida
  • PmHS2
  • Substrate specificity
  • UDP-GlcNAc
  • UDP-sugars

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology
  • Medicine(all)

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