title: EmbA is an essential arabinosyltransferase in Mycobacterium tuberculosis.
abstract: The Emb proteins (EmbA, EmbB, EmbC) are mycobacterial arabinosyltransferases involved in the biogenesis of the mycobacterial cell wall. EmbA and EmbB are predicted to work in unison as a heterodimer. EmbA and EmbB are involved in the formation of the crucial terminal hexaarabinoside motif [Arabeta(1-->2)Araalpha(1-->5)] [Arabeta(1-->2)Araalpha(1-->3)]Araalpha(1-->5)Araalpha1-->(Ara(6)) in the cell wall polysaccharide arabinogalactan. Studies conducted in Mycobacterium smegmatis revealed that mutants with disruptions in embA or embB are viable, although the growth rate was affected. In contrast, we demonstrate here that embA is an essential gene in Mycobacterium tuberculosis, since a deletion of the chromosomal gene could only be achieved when a second functional copy was provided on an integrated vector. Complementation of an embA mutant of M. smegmatis by M. tuberculosis embA confirmed that it encodes a functional arabinosyltransferase. We identified a promoter for M. tuberculosis embA located immediately upstream of the gene, indicating that it is expressed independently from the upstream gene, embC. Promoter activity from P(embA)((Mtb)) was sevenfold lower when assayed in M. smegmatis compared to M. tuberculosis, indicating that the latter is not a good host for genetic analysis of M. tuberculosis embA expression. P(embA)((Mtb)) activity remained constant throughout growth phases and after stress treatment, although it was reduced during hypoxia-induced non-replicating persistence. Ethambutol exposure had no effect on P(embA)((Mtb)) activity. These data demonstrate that M. tuberculosis embA encodes a functional arabinosyltransferase which is constitutively expressed and plays a critical role in M. tuberculosis.
authors.affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, CO 80523, USA.
authors.forename: Anita G
authors.initials: AG
authors.lastname: Amin
authors.affiliation: Centre for Infectious Disease, Barts and the London, Queen Mary's School of Medicine and Dentistry, London E1 2AT, UK.
authors.forename: Renan
authors.initials: R
authors.lastname: Goude
authors.affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, CO 80523, USA.
authors.forename: Libin
authors.initials: L
authors.lastname: Shi
authors.affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, CO 80523, USA.
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, CO 80523, USA.
authors.forename: Delphi
authors.initials: D
authors.lastname: Chatterjee
authors.affiliation: Centre for Infectious Disease, Barts and the London, Queen Mary's School of Medicine and Dentistry, London E1 2AT, UK.
authors.forename: Tanya
authors.initials: T
authors.lastname: Parish
mesh_terms: D019677:Artificial Gene Fusion; D004977:Ethambutol / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D005798:Genes, Bacterial; D020043:Genes, Essential; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D020102:Mycobacterium smegmatis / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D010430:Pentosyltransferases / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
chemical_list: D004977:Ethambutol; D010430:Pentosyltransferases; C101967:arabinosyltransferase; D001616:beta-Galactosidase