Modeling and Mutational analysis of Anion transporter 1 protein of Arabidopsis thaliana
2010 (English)In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 277, no Suppl. 1, 231-231 p.Article in journal, Meeting abstract (Other academic) Published
The thylakoid anion transporter 1 (ANTR1) from Arabidopsisthaliana, has been characterized as a Na-dependent Pi transporter when expressed in E. coli (1), but no data is yet available for the protein structure and amino acids involved in transport of Pi. In this study a three-dimensional structural model of ANTR1 was constructed in silico using the crystal structure of glycerol-3- phosphate/phosphate antiporter from E. coli as a template. Based on Multiple Sequence Alignments (MSAs) with other plant ANT- Rs and mammalian SLC17 homologues, ﬁve highly conserved amino acids involved in Pi transport have been identiﬁed, namely Arg-120, Ser-124 and Arg-201 inside the putative translocation pathway, Arg-228 and Asp-382 exposed at the cytoplasmic sur- face of the protein. The activity of the protein as a Na-dependent Pi transporter in the wild type and mutants was analyzed by het- erologous expression and uptake of radioactive Pi into E. coli cells. Substitution of the three Arg (120, 201 and 228) for Glu residues and of Asp-382 for an Asn residue resulted in an inac- tive ANTR1 transporter. All other mutants had sufﬁcient activity to allow measurement of kinetic parameters, attesting that the mutated proteins were functional. Based on our results, we pro- pose that Arg-201 is a critical residue for substrate binding and translocation, whereas Ser-124 may function as periplasmic gate- way for Na+ ions. Residue Arg-120 plays an important role in Pi binding and associated conformational changes, and ﬁnally that Arg-228 and Asp-382 only weakly participate in interactions allowing conformational changes to occur at the cytoplasmic sur-face of the transporter.
Place, publisher, year, edition, pages
Wiley-Blackwell, 2010. Vol. 277, no Suppl. 1, 231-231 p.
National CategoryEngineering and Technology
IdentifiersURN: urn:nbn:se:liu:diva-58961ISI: 000278565100804OAI: oai:DiVA.org:liu-58961DiVA: diva2:347780
35th Congress of the Federation-of-European-Biochemical-Societies, Gothenburg, Sweden, June 26-July 01, 2010