Insights into the structural determinants for specificity and transport by UreA, the specific urea transporter of Aspergillus

Ref ID: 18469

Author:

Lic Manuel Sanguinetti, Sotiris Amillis, Dr Claudio Scazzocchio, Sergio Pantano, Ana Ramón

Author address:

Sección Bioquí­mica, Departamento de Biologí­a Celular y Molecular, Facultad de Ciencias, UdelaR, Montevideo,
Uruguay Departamento de Botánica, Facultad de Biologí­a, Universidad de Atenas, Grecia Grupo de
Biosimulaciones, Institut Pasteur de Montevid

Full conference title:

11 th European Conference on Fungal Genetics

Abstract:

UreA, is a high8208;affinity urea/H+ symporter which seems to be the sole transport system specific for urea
inAspergillus nidulans. Homologous proteins have been characterized in Saccharomyces cerevisiae, Paxillus
involutus, Candida albicans, in the model plant Arabidopsis thaliana and more recently in Oryza
sativa (rice).Paralogues are present in the genomic sequences of all Aspergilli. The protein is predicted to consist of
15 transmembrane helical domains (TMSs), with an extracellular N8208;terminus and an intracellular C8208;terminus.
Little is known about the structure8208;function relationship of these membrane proteins. In order to address this
subject, we designed a mutational strategy based on 3D homology modelling of UreA and the identification of
conserved residues in all known fungal urea transporters. The functionality of the mutant proteins was assayed by
growth tests on urea and resistance to its toxic analogue, thiourea. All mutations were introduced on an UreA::GFP
fusion construct, which allowed us to follow the sub8208;cellular localization of mutant fusion proteins. This strategy
allowed us to identify a number of key residues involved in the recognition and/or translocation of urea across the
membrane. These mutations localize in helixes number 3, 7 or 11 which, according to homology modelling, are
predicted to be part of the substrate binding domain. A chemical mutagenesis approach has been also undertaken
which allowed for the identification of key residues for the functionality of the protein.
Our work constitutes the first mutational analysis in this family of transporters, providing insights into urea
transporters functionality.

Abstract Number: PR1.61

Conference Year: 2012

Link to conference website: http://www.ecfg.info/images/Abstract_Book_Electronic.pdf

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