dc.contributor.author | Gutsche, J | |
dc.contributor.author | Remminghorst, U | |
dc.contributor.author | Rehm, BHA | |
dc.date.accessioned | 2018-03-13T03:03:40Z | |
dc.date.available | 2018-03-13T03:03:40Z | |
dc.date.issued | 2006 | |
dc.identifier.issn | 0300-9084 | |
dc.identifier.doi | 10.1016/j.biochi.2005.06.003 | |
dc.identifier.uri | http://hdl.handle.net/10072/371079 | |
dc.description.abstract | AlgX was found to be an essential protein for alginate biosynthesis, but its function is unknown. In this study, an isogenic, marker-free algX-knock out mutant was generated. In-frame fusions of algX with phoA and lacZ were analysed, respectively. No LacZ-activity was detected, but the PhoA fusion showed alkaline phosphatase activity. These data indicated that the C-terminus of AlgX is located in the periplasm, but is not required for protein function. Accordingly, AlgX with C-terminal fusion of strep tag II restored alginate production in the algX-negative mutant and was purified under native conditions from periplasmic and crude cell extracts, respectively. AlgX was identified by MALDI/TOF-MS analysis of tryptic peptides. TritonX-100 mediated solubilisation of cytoplasmic membrane and subsequent strep tag II affinity chromatography led to purification of an AlgX-MucD (AlgY) protein complex as identified by MALDI/TOF-MS analysis. This data suggested a protein–protein interaction between AlgX and MucD (AlgY) with a 1:1 stoichiometry. Thus AlgX might exert its function via interaction with MucD (AlgY). Immunoelectron microscopic localisation of AlgX-strep tag II suggested a localisation close to the cytoplasmic membrane. | |
dc.description.peerreviewed | Yes | |
dc.language | English | |
dc.language.iso | eng | |
dc.publisher | Elsevier | |
dc.relation.ispartofpagefrom | 245 | |
dc.relation.ispartofpageto | 251 | |
dc.relation.ispartofissue | 3-4 | |
dc.relation.ispartofjournal | Biochimie | |
dc.relation.ispartofvolume | 88 | |
dc.subject.fieldofresearch | Biochemistry and cell biology | |
dc.subject.fieldofresearch | Biochemistry and cell biology not elsewhere classified | |
dc.subject.fieldofresearchcode | 3101 | |
dc.subject.fieldofresearchcode | 310199 | |
dc.title | Biochemical analysis of alginate biosynthesis protein AlgX from Pseudomonas aeruginosa: purification of an AlgX-MucD (AlgY) protein complex | |
dc.type | Journal article | |
dc.type.description | C1 - Articles | |
dc.type.code | C - Journal Articles | |
gro.hasfulltext | No Full Text | |
gro.griffith.author | Rehm, Bernd | |