INTRODUCTION: SELENOPROTEINS 1. Definition of selenoprotein Selenoproteins are Selenocysteine (Sec) containing proteins which have been identified in each of the 3 domains of life: eubacteria, archaea and eukaryotes. Sec, known as the 21st amino acid (aa), is encoded in these proteins by the codon UGA. Although it is normally a termination codon of protein synthesis, it is also used as a Sec codon [6][7]. Sec is an analog of cysteine (Cys), with the sulfur (S) atom present in the side chain of Cys being replaced by a Selenium (Se) atom. Se is an essential micronutrient which deficiency may lead to disease, otherwise in excessive amounts is recognized as toxic. Se is found in cells mostly in selenoproteins involved in redox systems and may have antioxidant protection capability. Figure 1. Structure of Selenocysteine. At the physiologic pH, its selenol is mostly deprotonated. Image extracted from Wikipedia. 2. Biosynthesis of selenoproteins The main form of Selenium in mammals is found as selenoproteins, that consist in the incorporation of selenocysteine (Sec or U) in their structure. The 21st amino acid is similar to cysteine only changing the Sulphur atom by Selenium, by this fact, selenocysteine could be thougth to derive from cysteine but it is really synthesised from serine [4]. Sec is codified as UGA codon in the genome, which at the same time is also a termination codon [5]. Selenocysteine has not a permanent pool in cells due to two major reasons: the first one is because of the risk of misincorporation of the amino acid in proteins replacing cysteine. This is dangerous for the cell because selenocysteine is more reactive compared to normal cysteine [6]. Second, selenite (a substrate to synthesise the amino acid) is highly reactive with oxygen and thioredoxin reductase, this leads to quick oxidation of NADPH and formation of reactive oxygen species [6]. 2.1 Synthesis and incorporation of selenocysteine in proteins The main source of Selenium is through Selenite and selenate, which are absorbed from aliments. Selenite, now in the body, is reduced to selenide (HSe-) by glutathione-glutaredoxin and thioredoxin systems [6]. Then, via catalysis by the selenoprotein SPS2 (selenophosphate synthetase 2), the selenide is converted to monoselenophosphate (H2SePO3-), which is the active selenium donor in the conversion from seryl-tRNAsec to Sec-tRNAsec (tRNA sepcific for selenocysteine). Presence of Seryl-tRNASec means that serine is loaded into Sec-tRNASec, from this scaffold the selenocysteine will be built and later incorporated onto the protein [6][7]. The first step is the phosphorylation of serine by Phosphoseryl tRNA kinase (PSTK) [5][7]. Afterwards, phosphoserine attached in the tRNA loses its phosphate and gets ligated the Selenium atom in its structure by the action of Selenocysteine synthase [6]. To incorporate selenocysteine into the protein, first of all it is necessary to correctly interpret the UGA codon as Sec instead of a termination codon and also the ribosome has to recognise the specific Sec-tRNASec carrying selenocysteine. To complete this, mRNA has a SECIS element (selenocysteine insertion sequence) that consists in a tertiary RNA structure similar to a hairpin in the 3’ UTR RNA end (in eukarya).[7] This will be recognised by several Binding Proteins as SBP2(SECIS Binding Protein 2), eEFSec (Selenocysteine Especific Elongation factor), Ribosomal protein L30, Sec43bp and SecS (SLA/LP) also known as Eukaryotic Selenocysteine Syntase) that will bind to Sec-tRNASec and will drive it towards the SECIS structure to add the amino acid into protein sequence [6][7]. Figure 2. Selenocysteine biosynthesis pathways in eukarya [7] . 3. Evolution of selenoproteins Regarding the evolutionary history of selenoproteins, it has been found that selenoproteins are present in all live domains. Further research on model animals revealed that selenoproteins sets are wide different among them. For example, green algae shows more than 20 selenoproteins in its genome while red algae, insects and nematodes display less than five. If we look at higher plants, only on American cranberries (known as Vaccinium macrocarpon [8]), has been detected the presence of the machinery necessary for synthesise selenocysteine and there has not been reported any on fungi. Other studies show how aquatic selenoproteomes are larger than the ones found in terrestrian animals, who tend to minimise them. This unique selenoproteins found only in aquatic species were generated by duplication as in bony fishes and it was also observed that some of them were lost across lineages by the replacement of selenocysteine by cysteine. This two facts together show how easily can selenoproteins change [9]. Analysing vertebrates, 21 selenoproteins were found in all of them: GPx1-4, TR1, TR3, Dio1, Dio2, Dio3, SelH, SelI, SelK, SelM, SelN, SelO, SelP, MsrB1 (methionine-R-sulfoxide reductase 1), SelS, SelT1, SelW1, Sep15 [9]. GPx1b, GPx3b, GPx4b, Dio3b, SelT2, MsrB1b and SelU1c were originated by the duplication of the genome in bony fishes. Additionally, some gene duplications were observed only in specific lineages of bony fishes. In zebrafish only, we found additional copies of SelO, SelT1 and SelW2, named respectively SelO2, SelT1b, and SelW2b [9]. After the split with fishes, several selenoproteins were generated also in the lineage to mammals. This is consistent with the idea that mammals reduced their utilization of Sec compared with fishes [8][9]. All the proteins will be studied with more detail in the discussion. Figure 3. Composition and Evolution of the Vertebrate and Mammalian Selenoproteomes [8] .