High-throughput method for in process monitoring of 3-O-sulfotransferase catalyzed sulfonation in bioengineered heparin synthesis

Authors
Lin, Lei
Yu, Yanlei
Zhang, Fuming
Zhang, Xing
Linhardt, Robert J.
ORCID
https://orcid.org/0000-0003-2219-5833
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Other Contributors
Issue Date
2019-12-01
Keywords
Biology , Chemistry and chemical biology , Chemical and biological engineering , Biomedical engineering
Degree
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Full Citation
High-throughput method for in process monitoring of 3-O-sulfotransferase catalyzed sulfonation in bioengineered heparin synthesis, L. Lin, Y. Yu, F. Zhang, X. Zhang, R. J. Linhardt, Analytical Biochemistry, 586, 113419, 2019.
Abstract
Bioengineered heparin (BEH) offers a potential alternative for the preparation of a safer pharmacological heparin. Construction of in-process control assays for tracking each enzymatic step during bioengineered heparin synthesis remains a challenge. Here, we report a high-throughput sensing platform based on enzyme-linked immunosorbent assay (ELISA) and enzymatic signal amplification that allows the rapid and accurate monitoring of the 3-OST sulfonation in BEH synthesis process. The anticoagulant activity of target BEH was measured to reflect the degree of sulfonation by testing its competitive antithrombin (AT) binding ability. BEH samples with different sulfonation degrees show different AT protein binding capacity and thus changes the UV response to a different extent. This BEH-induced signal can be conveniently and sensitively monitored by the plate sensing system, which benefits from its high sensitivity brought in by the enzymatic signal amplification. Furthermore, modification convenience and mechanical robustness also ensure the stability of the test platform. This proposed strategy exhibits excellent analytical performance in both BEH activity analysis and 3-OST sulfonation evaluation. The simple and sensitive plate system shows great potential in developing on-chip, high-throughput methods for fundamental biochemical process research, drug discovery, and clinic diagnostics.
Description
Analytical Biochemistry, 586, 113419
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Department
The Linhardt Research Labs.
The Shirley Ann Jackson, Ph.D. Center for Biotechnology and Interdisciplinary Studies (CBIS)
Publisher
Relationships
The Linhardt Research Labs Online Collection
Rensselaer Polytechnic Institute, Troy, NY
Analytical Biochemistry
https://harc.rpi.edu/
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