Deciphering a pathway of Halobacterium salinarum N‐glycosylation

Genomic analysis points to N‐glycosylation as being a common posttranslational modification in Archaea. To date, however, pathways of archaeal N‐glycosylation have only been described for few species. With this in mind, the similarities of N‐linked glycans decorating glycoproteins in the haloarchaea Haloferax volcanii and Halobacterium salinarum directed a series of bioinformatics, genetic, and biochemical experiments designed to describe that Hbt. salinarum pathway responsible for biogenesis of one of the two N‐linked oligosaccharides described in this species. As in Hfx. volcanii , where agl ( a rchaeal gl ycosylation) genes that encode proteins responsible for the assembly and attachment of a pentasaccharide to target protein Asn residues are clustered in the genome, Hbt. salinarum also contains a group of clustered homologous genes ( VNG 1048G ‐ VNG 1068G ). Introduction of these Hbt. salinarum genes into Hfx. volcanii mutant strains deleted of the homologous sequence restored the lost activity. Moreover, transcription of the Hbt. salinarum genes in the native host, as well as in vitro biochemical confirmation of the predicted functions of several of the products of these genes provided further support for assignments made following bioinformatics and genetic experiments. Based on the results obtained in this study, the first description of an N‐glycosylation pathway in Hbt. salinarum is offered.