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    Biochemical characterization of the vacuolar palmitoyl acyltransferase (2003)

    Art
    Zeitschriftenartikel / wissenschaftlicher Beitrag
    Autoren
    Veit, Michael
    Dietrich, Lars E P
    Ungermann, Christian
    Quelle
    FEBS letters; 540(1/33) — S. 101–105
    ISSN: 0014-5793
    Sprache
    Englisch
    Verweise
    Pubmed: 12681491
    Kontakt
    Institut für Immunologie

    Robert-von-Ostertag-Str. 7-13
    Gebäude 35
    14163 Berlin
    +49 30 838 51834

    Abstract / Zusammenfassung

    Vacuole fusion requires Sec18p-dependent acylation of the armadillo-repeat protein Vac8p that has been isolated with cis-SNARE complexes. To gain more insight into the mechanism of acylation, we analyzed the palmitoylation reaction on isolated vacuoles or in vacuolar detergent extracts. Recombinant Vac8p is palmitoylated when added to vacuoles and is anchored to membranes after modification. The palmitoyl acyltransferase (PAT) extracted from vacuolar membranes is functional in detergent extracts and shows all characteristics of an enzymatic activity: It modifies exogenous Vac8p in a temperature-, dose- and time-dependent manner, and is sensitive to bromo-palmitate, a known inhibitor of protein palmitoylation in vivo. Importantly, PAT is specific for palmitoyl-CoA, since myristoyl- and stearyl-CoA can compete with labeled Pal-CoA only at 10-fold higher amounts.