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Recombinant canine single chain insulin analogues: Insulin receptor binding capacity and ability to stimulate glucose uptake

Adams, J P and Holder, A L and Catchpole, B (2014) Recombinant canine single chain insulin analogues: Insulin receptor binding capacity and ability to stimulate glucose uptake. VETERINARY JOURNAL, 202 (3). pp. 436-42.

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Virtually all diabetic dogs require exogenous insulin therapy to control their hyperglycaemia. In the UK, the only licensed insulin product currently available is a purified porcine insulin preparation. Recombinant insulin is somewhat problematic in terms of its manufacture, since the gene product (preproinsulin) undergoes substantial post-translational modification in pancreatic β cells before it becomes biologically active. The aim of the present study was to develop recombinant canine single chain insulin (SCI) analogues that could be produced in a prokaryotic expression system and which would require minimal processing. Three recombinant SCI constructs were developed in a prokaryotic expression vector, by replacing the insulin C-peptide sequence with one encoding a synthetic peptide (GGGPGKR), or with one of two insulin-like growth factor (IGF)-2 C-peptide coding sequences (human: SRVSRRSR; canine: SRVTRRSSR). Recombinant proteins were expressed in the periplasmic fraction of Escherichia coli and assessed for their ability to bind to the insulin and IGF-1 receptors, and to stimulate glucose uptake in 3T3-L1 adipocytes. All three recombinant SCI analogues demonstrated preferential binding to the insulin receptor compared to the IGF-1 receptor, with increased binding compared to recombinant canine proinsulin. The recombinant SCI analogues stimulated glucose uptake in 3T3-L1 adipocytes compared to negligible uptake using recombinant canine proinsulin, with the canine insulin/cIGF-2 chimaeric SCI analogue demonstrating the greatest effect. Thus, biologically-active recombinant canine SCI analogues can be produced relatively easily in bacteria, which could potentially be used for treatment of diabetic dogs.

Item Type: Article
RVC Publication Type: Research (full) paper
WoS ID: 000348017500007
DOI: https://doi.org/10.1016/j.tvjl.2014.09.027
Departments: Pathobiology and Population Sciences
Research Programmes: Comparative Physiology & Medicine > Immune Regulation and Cancer
Depositing User: RVC Auto-import
Date Deposited: 08 Jul 2015 05:01
Last Modified: 31 May 2018 12:51
URI: http://researchonline.rvc.ac.uk/id/eprint/9400