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dc.contributor.authorRemminghorst, Uwe
dc.contributor.authorHay, Iain D
dc.contributor.authorRehm, Bernd HA
dc.date.accessioned2018-07-05T12:30:30Z
dc.date.available2018-07-05T12:30:30Z
dc.date.issued2009
dc.identifier.issn0168-1656
dc.identifier.doi10.1016/j.jbiotec.2009.02.006
dc.identifier.urihttp://hdl.handle.net/10072/369678
dc.description.abstractThe topology of Alg8, the proposed catalytic subunit of the alginate polymerase, was assessed using PhoA and LacZ fusion protein analysis. This analysis suggested that the periplasmic loop comprises only three amino acid residues with the adjacent transmembrane helices at positions 361–387 and 393–416. Accordingly, the extended cytosolic loop could be located at positions 71–361 and was proposed to contain important catalytic residues. Further experimental evidence for this cytosolic domain was obtained by independently demonstrating this protein region as purified soluble protein domain. The soluble protein domain was identified by MALDI-TOF/MS and presumably represents the cytosolic catalytic domain of Alg8. Site-directed mutagenesis of 11 conserved residues in the cytosolic loop showed that D-188/D-190 (DXD motif), D-295/D-296 (acid–base catalysts) and K-297 were each essential for in vivo polymerase activity, whereas D-179/D-181 (DXD motif), C-244, R-263, D-279, and E-282 were not directly involved in the polymerisation reaction. The role of these amino acid residues with respect to the catalysed alginate polymerisation reaction was discussed with the aid of the recently developed structural model of Alg8.
dc.description.peerreviewedYes
dc.languageEnglish
dc.language.isoeng
dc.publisherElsevier
dc.relation.ispartofpagefrom176
dc.relation.ispartofpageto183
dc.relation.ispartofissue3-4
dc.relation.ispartofjournalJournal of Biotechnology
dc.relation.ispartofvolume140
dc.subject.fieldofresearchBiochemistry and Cell Biology not elsewhere classified
dc.subject.fieldofresearchBiological Sciences
dc.subject.fieldofresearchEngineering
dc.subject.fieldofresearchTechnology
dc.subject.fieldofresearchcode060199
dc.subject.fieldofresearchcode06
dc.subject.fieldofresearchcode09
dc.subject.fieldofresearchcode10
dc.titleMolecular characterization of Alg8, a putative glycosyltransferase, involved in alginate polymerisation
dc.typeJournal article
dc.type.descriptionC1 - Articles
dc.type.codeC - Journal Articles
gro.hasfulltextNo Full Text
gro.griffith.authorRehm, Bernd


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