Membrane topology of aspartate:alanine antiporter AspT from Comamonas testosteroni

Takashi Fujiki, Kei Nanatani, Kei Nishitani, Kyoko Yagi, Fumito Ohnishi, Hiroshi Yoneyama, Takafumi Uchida, Tasuku Nakajima, Keietsu Abea

    Research output: Contribution to journalArticlepeer-review

    4 Citations (Scopus)

    Abstract

    We cloned the aspT gene encoding the L-aspartate:L-alanine antiporter AspTCt in Comamonas testosteroni genomic DNA. Analysis of the nucleotide sequence revealed that C. testosteroni has an asp operon containing aspT upstream of the L-aspartate 4-decarboxylase gene, and that the gene order of the asp operon of C. testosteroni is the inverse of that of Tetragenococcus halophilus. We used proteoliposomes to confirm the transport processes of AspTCt. To elucidate the two-dimensional structure of AspT Ct, we analysed its membrane topology by means of alkaline phosphatase (PhoA) and β-lactamase (BlaM) fusion methods. The fusion analyses revealed that AspTCt has seven transmembrane segments (TMs), a large cytoplasmic loop containing ∼200 amino acid residues between TM4 and TM5, a cytoplasmic N-terminus, and a periplasmic C-terminus. These results suggest that the orientation of the N-terminus of AspTCt differs from that of tetragenococcal AspT, even though these two AspT orthologues catalyse the same transport reactions.

    Original languageEnglish
    Pages (from-to)85-91
    Number of pages7
    JournalJournal of biochemistry
    Volume141
    Issue number1
    DOIs
    Publication statusPublished - 2007 Jan 1

    Keywords

    • Aspartate:alanine antiporter
    • Bacteria
    • Bioenergetics
    • Fusion methods
    • Membrane topology

    ASJC Scopus subject areas

    • Biochemistry
    • Molecular Biology

    Fingerprint

    Dive into the research topics of 'Membrane topology of aspartate:alanine antiporter AspT from Comamonas testosteroni'. Together they form a unique fingerprint.

    Cite this