High-yield production of apoplast-directed human adenosine deaminase in transgenic tobacco BY-2 cell suspensions

Article


Singhabahu, Sanjeewa, George, John and Bringloe, D. 2015. High-yield production of apoplast-directed human adenosine deaminase in transgenic tobacco BY-2 cell suspensions. Biotechnology and Applied Biochemistry. 62 (1), pp. 87-93.
AuthorsSinghabahu, Sanjeewa, George, John and Bringloe, D.
Abstract

Adenosine deaminase (ADA) deficiency, where a deleterious mutation in the ADA gene of patients results in a dysfunctional immune system, is ultimately caused by an absence of ADA. Over the last 25 years the disease has been treated with PEG-ADA, made from purified bovine ADA coupled with polyethylene glycol (PEG). However, it is thought that an enzyme replacement therapy protocol based on recombinant human ADA would probably be a more effective treatment. With this end in mind, a human ADA cDNA was inserted into plant expression vectors used to transform tobacco plant cell suspensions. Transgenic calli expressing constructs containing apoplast-directing signals showed significantly higher levels of recombinant ADA expression than calli transformed with cytosolic constructs. The most significant ADA activities, however, were measured in the media of transgenic cell suspensions prepared from high expressing transformed calli: where incorporation of a signal for arabinogalactan addition to ADA led to a recombinant protein yield of approximately 16 mg L−1, a 336-fold increase over ADA produced by cell suspensions transformed with a cytosolic construct.

Keywordsadenosine deaminase; apoplast; molecular pharming; plant cell culture; recombinant protein; tobacco
JournalBiotechnology and Applied Biochemistry
Journal citation62 (1), pp. 87-93
ISSN0885-4513
1470-8744
Year2015
PublisherWiley for International Union of Biochemistry and Molecular Biology (IUBMB)
Digital Object Identifier (DOI)doi:10.1002/bab.1240
Publication dates
Print13 Feb 2015
Publication process dates
Deposited08 Feb 2017
Accepted03 May 2014
FunderUniversity of East London
University of East London
Copyright informationThis is the peer reviewed version of the following article: Singhabahu, S., George, J. and Bringloe, D. (2015), High-yield production of apoplast-directed human adenosine deaminase in transgenic tobacco BY-2 cell suspensions. Biotechnology and Applied Biochemistry, 62: 87–93, which has been published in final form at doi:10.1002/bab.1240. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.
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