SECIS element variability and its role in selenocysteine versus cysteine utilization in SelD enzymes

Abstract

Selenophosphate synthetase SelD from Haemophilus influenzae (HinSelD) has a selenocysteine (Sec) residue at its active site, while its Escherichia coli homolog (EcoSelD), which shares 65% amino acid sequence identity, contains cysteine (Cys) instead. This difference prompts questions about the evolutionary divergence between Sec-type and Cys-type SelD enzymes. We used bioinformatics tools to compare the selenocysteine insertion sequence (SECIS) elements of the Sec-type selD gene with the corresponding sequence regions of the Cys-type genes. Our analysis showed vital conservation between the UGA Sec codon and SECIS secondary structures. We also tested if HinSelD SECIS could support Sec insertion in E. coli. Results indicated that HinSelD SECIS is recognized by E. coli, enabling Sec incorporation. Nucleotide differences between HinSelD and EcoSelD SECIS regions affected translation efficiency, with mutants G69A, A75G, G77A, and U84C showing 93%, 81%, 69%, and 69% of wild-type translation levels, respectively. Additionally, the Sec16Cys mutant of HinSelD exhibited a similar expression level compared to the wild-type, suggesting the secondary structure of the SECIS does not inhibit the translation of the preceding UGC codon for Cys.

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