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dc.contributor.authorTavoularis, S. N.en_US
dc.contributor.authorTazebay, U. H.en_US
dc.contributor.authorDiallinas, G.en_US
dc.contributor.authorSideridou, M.en_US
dc.contributor.authorRosa, A.en_US
dc.contributor.authorScazzocchio, C.en_US
dc.contributor.authorSophianopoulou, V.en_US
dc.date.accessioned2016-02-08T10:29:13Z
dc.date.available2016-02-08T10:29:13Z
dc.date.issued2003en_US
dc.identifier.issn0968-7688
dc.identifier.urihttp://hdl.handle.net/11693/24427
dc.description.abstractPrnB, the L-proline transporter of Aspergillus nidulans, belongs to the Amino acid Polyamine Organocation (APC) transporter family conserved in prokaryotes and eukaryotes. In silico analysis and limited biochemical evidence suggest that APC transporters comprise 12 transmembrane segments (TMS) connected with relatively short hydrophilic loops (L). However, very little is known on the structure-function relationships in APC transporters. This work makes use of the A. nidulans PrnB transporter to address structure-function relationships by selecting, constructing and analysing several prnB mutations. In the sample, most isolated missense mutations affecting PrnB function map in the borders of cytoplasmic loops with transmembrane domains. These are I119N and G120W in L2-TMS3, F278V in L6-TMS7, NRT378NRTNRT and PY382PYPY in L8-TMS9 and T456N in L10-TMS11. A single mutation (G403E) causing, however, a very weak phenotype, maps in the borders of an extracellular loop (L9-TMS10). An important role of helix TMS6 for proline binding and transport is supported by mutations K245L and, especially, F248L that clearly affect PrnB uptake kinetics. The critical role of these residues in proline binding and transport is further shown by constructing and analysing isogenic strains expressing selected prnB alleles fused to the gene encoding the Green Fluorescent Protein (GFP). It is shown that, while some prnB mutations affect proper translocation of PrnB in the membrane, at least two mutants, K245E and F248L, exhibit physiological cellular expression of PrnB and, thus, the corresponding mutations can be classified as mutations directly affecting proline binding and/or transport. Finally, comparison of these results with analogous studies strengthens conclusions concerning amino acid residues critical for function in APC transporters.en_US
dc.language.isoEnglishen_US
dc.source.titleMolecular Membrane Biologyen_US
dc.relation.isversionofhttp://dx.doi.org/10.1080/0968768031000106339en_US
dc.subjectAspergillus nidulansen_US
dc.subjectMutagenesisen_US
dc.subjectProlineen_US
dc.subjectStructure-function analysisen_US
dc.subjectTransporten_US
dc.subjectAmino aciden_US
dc.subjectCarrier proteinen_US
dc.subjectGreen fluorescent proteinen_US
dc.titleMutational analysis of the major proline transporter (PrnB) of aspergillus nidulansen_US
dc.typeArticleen_US
dc.departmentDepartment of Molecular Biology and Geneticsen_US
dc.citation.spage285en_US
dc.citation.epage297en_US
dc.citation.volumeNumber20en_US
dc.citation.issueNumber4en_US
dc.identifier.doi10.1080/0968768031000106339en_US
dc.publisherTaylor & Francisen_US


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