Issue 7, 2017

Converting Pasteurella multocida α2–3-sialyltransferase 1 (PmST1) to a regioselective α2–6-sialyltransferase by saturation mutagenesis and regioselective screening

Abstract

A microtiter plate-based screening assay capable of determining the activity and regioselectivity of sialyltransferases was developed. This assay was used to screen two single-site saturation libraries of Pasteurella multocida α2–3-sialyltransferase 1 (PmST1) for α2–6-sialyltransferase activity and total sialyltransferase activity. PmST1 double mutant P34H/M144L was found to be the most effective α2–6-sialyltransferase and displayed 50% reduced donor hydrolysis and 50-fold reduced sialidase activity compared to the wild-type PmST1. It retained the donor substrate promiscuity of the wild-type enzyme and was used in an efficient one-pot multienzyme (OPME) system to selectively catalyze the sialylation of the terminal galactose residue in a multigalactose-containing tetrasaccharide lacto-N-neotetraoside.

Graphical abstract: Converting Pasteurella multocida α2–3-sialyltransferase 1 (PmST1) to a regioselective α2–6-sialyltransferase by saturation mutagenesis and regioselective screening

Supplementary files

Article information

Article type
Paper
Submitted
11 Dec. 2016
Accepted
20 Janv. 2017
First published
20 Janv. 2017

Org. Biomol. Chem., 2017,15, 1700-1709

Converting Pasteurella multocida α2–3-sialyltransferase 1 (PmST1) to a regioselective α2–6-sialyltransferase by saturation mutagenesis and regioselective screening

J. B. McArthur, H. Yu, J. Zeng and X. Chen, Org. Biomol. Chem., 2017, 15, 1700 DOI: 10.1039/C6OB02702D

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