Binding affinity and capacity of putative adaptor-mediated sorting of a Type I membrane protein in Leishmania mexicana

Weise, Frank and Thilo, Lutz and Engstler, Markus and Wiese, Martin and Benzel, Isabel and Kühn, Christina and Bühring, Hans-Jörg and Overath, Peter (2005) Binding affinity and capacity of putative adaptor-mediated sorting of a Type I membrane protein in Leishmania mexicana. Molecular and Biochemical Parasitology, 142 (2). pp. 203-11. ISSN 0166-6851

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Abstract

The membrane-bound acid phosphatase (MBAP), a Type I membrane protein predominantly associated with endosomal/lysosomal structures of Leishmania mexicana promastigotes, contains motifs in its cytosolic COOH-terminal tail (-MEVWRRYMKFKNKQSEAIIV-COOH) akin to tyrosine- and di-leucine-based sorting signals in multicellular organisms. Here, we first show that the COOH-terminal residues IIV of MBAP, but not the Y-residue, are required for endosomal targeting, suggesting specific binding to an adaptor complex at the cell surface. We then determine whether specific binding can be saturated by analysing the efficiency of endosomal targeting for increasing numbers of MBAP molecules per cell. The ratio of the steady-state abundance of wild-type MBAP on the cell surface to MBAP on endosomes increases until the distribution is no longer different from that observed for a mutant MBAP which lacks the IIV-motif or for a glycosylphosphatidylinositol-anchored form, both of which are distributed according to bulk membrane flow. A quantitative analysis of these in vivo results indicates specific binding to a putative adaptor complex with an affinity of about 10-4M to 50,000 sorting sites on the cell surface.

ORCID iDs

Weise, Frank, Thilo, Lutz, Engstler, Markus, Wiese, Martin ORCID logoORCID: https://orcid.org/0000-0003-4493-0835, Benzel, Isabel, Kühn, Christina, Bühring, Hans-Jörg and Overath, Peter;