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Title 

Enhanced bacterial α(2,6)-sialyltransferase reaction through an inhibition of its inherent sialidase activity by dephosphorylation of cytidine-5'-monophosphate

Authors 

Ji-Yeon KangSe-Jong LimOh Suk KwonSeung Goo LeeH H KimDoo-Byoung Oh

Publisher 

Public Library of Science

Issue Date 

2015

Citation 

Plos One, vol. 10, no. 7, pp. e0133739-e0133739

Abstract 

Bacterial α(2,6)-sialyltransferases (STs) from Photobacterium damsela, Photobacterium sp. JT-ISH-224, and P. leiognathi JT-SHIZ-145 were recombinantly expressed in Escherichia coli and their ST activities were compared directly using a galactosylated bi-antennary N-glycan as an acceptor substrate. In all ST reactions, there was an increase of sialylated glycans at shorter reaction times and later a decrease in prolonged reactions, which is related with the inherent sialidase activities of bacterial STs. These sialidase activities are greatly increased by free cytidine monophosphate (CMP) generated from a donor substrate CMP-N-acetylneuraminic acid (CMP-Neu5Ac) during the ST reactions. The decrease of sialylated glycans in prolonged ST reaction was prevented through an inhibition of sialidase activity by simple treatment of alkaline phosphatase (AP), which dephosphorylates CMP to cytidine. Through supplemental additions of AP and CMP-Neu5Ac to the reaction using the recombinant α(2,6)-ST from P. leiognathi JT-SHIZ-145 (P145-ST), the content of bisialylated N-glycan increased up to ~98% without any decrease in prolonged reactions. This optimized P145-ST reaction was applied successfully for α(2,6)-sialylation of asialofetuin, and this resulted in a large increase in the populations of multi-sialylated N-glycans compared with the reaction without addition of AP and CMP-Neu5Ac. These results suggest that the optimized reaction using the recombinant P145-ST readily expressed from E. coli has a promise for economic glycan synthesis and glyco-conjugate remodeling.

ISSN 

1932-6203

Link 

http://dx.doi.org/10.1371/journal.pone.0133739

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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