A Pasteurella multocida sialyltransferase displaying dual trans-sialidase activities for production of 3′-sialyl and 6′-sialyl glycans

Yao Guo, Carsten Jers, Anne S. Meyer, Anis Arnous, Haiying Li, Finn Kirpekar, Jørn Dalgaard Mikkelsen

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

tThis study examined a recombinant Pasteurella multocida sialyltransferase exhibiting dual trans-sialidase activities. The enzyme catalyzed trans-sialylation using either 2-O-(p-nitrophenyl)--d-N-acetylneuraminic acid or casein glycomacropeptide (whey protein) as the sialyl donor and lactose asthe acceptor, resulting in production of both 3-sialyllactose and 6-sialyllactose. This is the first studyreporting -2,6-trans-sialidase activity of this sialyltransferase (EC 2.4.99.1 and 2.4.99.4). A responsesurface design was used to evaluate the effects of three reaction parameters (pH, temperature, andlactose concentration) on enzymatic production of 3- and 6-sialyllactoses using 5% (w/v) casein glyco-macropeptide (equivalent to 9 mM bound sialic acid) as the donor. The maximum yield of 3-sialyllactose(2.75 ± 0.35 mM) was achieved at a reaction condition with pH 6.4, 40◦C, 100 mM lactose after 6 h; andthe largest concentration of 6-sialyllactose (3.33 ± 0.38 mM) was achieved under a condition with pH5.4, 40◦C, 100 mM lactose after 8 h. 6-sialyllactose was presumably formed from -2,3 bound sialic acidin the casein glycomacropeptide as well as from 3-sialyllactose produced in the reaction. The kcat/Kmvalue for the enzyme using 3-sialyllactose as the donor for 6-sialyllactose synthesis at pH 5.4 and 40◦Cwas determined to be 23.22 ± 0.7 M−1s−1. Moreover, the enzyme was capable of catalyzing the synthesisof both 3- and 6-sialylated galactooligosaccharides, when galactooligosaccharides served as acceptors.
Original languageEnglish
JournalJournal of Biotechnology
Volume170
Pages (from-to)60-67
Number of pages8
ISSN0168-1656
DOIs
Publication statusPublished - 2014

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