Contaminants in Commercial Preparations of ‘Purified’ Small Leucine-Rich Proteoglycans May Distort Mechanistic Studies

Owen, SJ; Jones, PW; Caterson, B; Shirran, SL; Fuller, H; Botting, CH; Roberts, S

Abstract

This paper reports the perplexing results that came about because of seriously impure commercially available reagents. Commercial reagents and chemicals are routinely ordered by scientists and are expected to have been rigorously assessed for their purity. Unfortunately, we found this assumption to be risky. Extensive work was carried out within our laboratory using commercially-sourced preparations of the small leucine-rich proteoglycans, decorin and biglycan, to investigate their influence on nerve cell growth. Unusual results compelled us to analyse the composition and purity of both preparations of these proteoglycans using both mass spectrometry and Western blotting, with and without various enzymatic deglycosylations. Commercial ‘decorin’ and ‘biglycan’ were found to contain a mixture of proteoglycans including not only both decorin and biglycan but also fibromodulin and aggrecan. The unexpected effects of ‘decorin’ and ‘biglycan’ on nerve cell growth could be explained by these impurities. Decorin and biglycan contain either chondroitin or dermatan sulphate glycosaminoglycan chains whilst fibromodulin only contains keratan sulphate and the large (>2,500 kDa), highly glycosylated aggrecan, contains both keratan and chondroitin sulphate. The different structure, molecular weights and composition of these impurities significantly affected our work and any conclusions that could be made. These findings beg the question as to whether scientists need to verify the purity of each commercially obtained reagent used in their experiments. The implications of these findings are vast, since the effects of these impurities may already have led to inaccurate conclusions and reports in the literature with concomitant loss of researchers’ funds and time.

Citation

Owen, S., Jones, P., Caterson, B., Shirran, S., Fuller, H., Botting, C., & Roberts, S. (2016). Contaminants in Commercial Preparations of ‘Purified’ Small Leucine-Rich Proteoglycans May Distort Mechanistic Studies. Bioscience Reports, https://doi.org/10.1042/BSR20160465

• Acceptance Date: Dec 19, 2016
• Publication Date: Dec 19, 2016
• Journal: Bioscience Reports
• Print ISSN: 0144-8463
• Publisher: Portland Press
• DOI: https://doi.org/10.1042/BSR20160465
• Keywords: biglycan, commercial preparations, decorin, glycosaminoglycans, mass spectrometry, small leucine-rich proteoglycans
• Publisher URL: http://dx.doi.org/10.1042/BSR20160465