Conformational Design and Characterisation of a Truncated Diamine Oxidase from Arthrobacter globiformis

Conformational Design and Characterisation of a Truncated Diamine Oxidase from Arthrobacter globiformis

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High-Throughput 2018, 7(3), 21; https://doi.org/10.3390/ht7030021
Nur Nadia Razali 1,2 ,Nur Hafizah Hashim 2,3 ,Adam Thean Chor Leow 1,2,3 andAbu Bakar Salleh 1,2,
Laboratory of Molecular Biomedicine, Institute of Bioscience, Universiti Putra Malaysia, Serdang 43400, Malaysia
Enzyme and Microbial Technology Research Center, Faculty of Biotechnology and Biomolecular Science, Universiti Putra Malaysia, Serdang 43400, Malaysia
Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang 43400, Malaysia
Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang 43400, Malaysia

Abstract

A functional mini protein can be developed by miniaturising its size. The minimisation technique provides an excellent model system for studying native enzymes, especially in creating an alternative novel biocatalyst. Miniaturised proteins may have enhanced stability, a crucial characteristic for large-scale production and industrial applications. In this study, a huge enzyme molecule, known as diamine oxidase (DAO, comprising 700 amino acids), was selected to undergo the process.

Methods

By retaining the arrangement of the original functional sites of DAO in the fourth domain, a mini DAO can be designed via homology modelling. After several downsizing processes, a final configuration of 220 amino acids displayed high binding affinity towards histamine, a short-chain substrate that was catalysed by the parental DAO. The configuration also showed enhanced affinity towards a long-chain substrate known as spermidine. The gene for the designed protein was cloned and expressed in pET102/TOPO vector and overexpressed in E. coli BL21 (DE3).

Results

The new mini DAO had similar temperature tolerance and versatile substrates specificity characteristics as its parental protein. An active mini-protein with these characteristics is potentially useful for several applications such as detecting biogenic amines in the biological fluids and the environment that may give rise to health issues. 

RELATED INFO

Histamine in food

Several foods provoke histamine accumulation in the organism, but not all are necessarily rich in this amine. High intakes of foods rich in other amines such as putrescine,

Que es el Déficit de DAO
What is DAO Deficiency?

DAO Deficiency is an alteration in the metabolism of food histamine that occurs when the activity of the Diamine Oxidase (DAO) enzyme is reduced. That is, when for

Histamine in food

Several foods provoke histamine accumulation in the organism, but not all are necessarily rich in this amine. High intakes of foods rich in other amines such as putrescine,

Que es el Déficit de DAO
What is DAO Deficiency?

DAO Deficiency is an alteration in the metabolism of food histamine that occurs when the activity of the Diamine Oxidase (DAO) enzyme is reduced. That is, when for