Characterisation of selenoproteins in fish feeds and in farmed salmon

Selenium is an essential, but also potential toxic mineral that is present in high natural levels in marine fish feed. The current practice in the aquaculture industry towards increased replacement of marine ingredients with plant ingredients affects both the natural levels and the chemical forms of selenium in fish feeds. In organisms, the biological functions related to selenium are mediated by selenium-containing proteins, i.e. the selenoproteins. The number of selenoproteins varies between different animal species, and teleost fish such as Atlantic salmon (Salmo salar) feature a higher number of selenoproteins compared to vertebrates. The selenoproteome of Atlantic salmon (Salmo salar) has, however, not been extensively studied. The present project focus on the characterization of selenoproteins in farmed Atlantic, and to further study possible effects on selenoproteins when salmon is fed different dietary sources of selenium. For the comprehensive characterisation of selenoproteins, suitable analytical workflows need to be developed. To analyse selenoproteins in fish feed and salmon a HPLC (high pressure liquid chromatography) coupled to an ICP-MS (inductively coupled plasma mass spectrometry) for selective detection of selenium. Using this methodology one can study the selenoprotein separated by size, and to compare the selenoprotein profile in fish feed and salmon tissue. Also, a methodology based on analysis of the selenoamino acids selenomethionine (SeMet) and selenocysteine (SeCys) has been developed for feeds and salmon muscle tissue. The results show that salmon fed SeMet-yeast contained SeMet as the major selenium compound in the muscle tissue, whereas salmon given feed containing inorganic selenium contained lower amount of SeMet. The results suggest that selenium is mainly bound unspecific to proteins in muscle tissue of salmon when the fish has fed SeMet-yeast dietary source. In this project also a combination of proteomics and trace element speciation techniques are used. In the course of the present project, we have developed a method for the determination of selenoproteins in salmon liver tissues. Using HPLC coupled to both an ICP-MS and a HR-MS (high resolution time of flight mass spectrometry) we are able to separate and detect selenium containing peptides and to simultaneously identify the molecular structures of the peptides. The analytical work is performed in collaboration with the Trace Element Speciation Laboratory at University of Aberdeen, Scotland. To further identify the proteins from the analytical data, database searches are commonly used. However, due to technical bioinformatics challenges in general, only few selenoproteins are listed in current versions of protein databases. This lack of data in the past has hampered any efforts to fully characterise the Atlantic salmon selenoproteome. The publication of the fully annotated Atlantic salmon genome, however, makes it possible to predict the salmon selenoproteins using bioinformatics tools. In collaboration with bioinformatics scientists from Harvard Medical School in Boston, USA, it has been predicted a comprehensive set of salmon selenoproteins. Currently we are in the process of experimentally validating the in silico predictions using the hyphenated analytical tools we developed earlier in the project. By combining the theoretical predictions with the experimental mass spectrometry data, we aim to obtain a comprehensive phenotypic description of the salmon selenoproteome. Based the salmon selenoproteome, in the future, we also will be able to assess how changes of concentrations and species of selenium in fish feeds affects the abundance and function selenoproteins.

Fish feeds contain high natural levels of the essential, but also potential toxic trace element selenium (Se). The current practice in the aquaculture industry towards increased replacement of marine ingredients with plant ingredients affects both the natural Se levels and the chemical forms of Se in the feeds. In organisms, the biological functions related to Se are mediated by the expression of selenium-bound proteins, i.e. the selenoproteins. Little is known about the bioavailability of Se in marine-based feed compared to plant-based feed, and how supplementation of Se (e.g. in the form of Se-enriched yeast or other Se-sources) in feeds affect the expression of selenoproteins in farmed fish. The present project will apply novel analytical approaches based on proteomics and advanced trace element analysis for characterising and identifying selenoproteins in fish feed and in farmed Atlantic salmon fed different dietary sources of Se. To date there are few studies on the selenoproteome in Atlantic salmon, and through this project valuable information on how the chemical forms of Se in feeds affect the bioavailability of Se and the selenoprotein expression in fish, will be obtained.