Structure of palmitoylated BET3: insights into TRAPP complex assembly and membrane localization.
BET3 is a component of TRAPP, a complex involved in the tethering of transport vesicles to the cis-Golgi membrane. The crystal structure of human BET3 has been determined to 1.55-A resolution. BET3 adopts an alpha/beta-plait fold and forms dimers in the crystal and in solution, which predetermines the architecture of TRAPP where subunits are present in equimolar stoichiometry. A hydrophobic pocket within BET3 buries a palmitate bound through a thioester linkage to cysteine 68. BET3 and yeast Bet3p are palmitoylated in recombinant yeast cells, the mutant proteins BET3 C68S and Bet3p C80S remain unmodified. Both BET3 and BET3 C68S are found in membrane and cytosolic fractions of these cells; in membrane extractions, they behave like tightly membrane-associated proteins. In a deletion strain, both Bet3p and Bet3p C80S rescue cell viability. Thus, palmitoylation is neither required for viability nor sufficient for membrane association of BET3, which may depend on protein-protein contacts within TRAPP or additional, yet unidentified modifications of BET3. A conformational change may facilitate palmitoyl extrusion from BET3 and allow the fatty acid chain to engage in intermolecular hydrophobic interactions.
Turnbull
Andrew P
AP
Max-Delbrück-Centrum für Molekulare Medizin, Berlin, Germany.
Kümmel
Daniel
D
Prinz
Bianka
B
Holz
Caterina
C
Schultchen
Jeffrey
J
Lang
Christine
C
Niesen
Frank H
FH
Hofmann
Klaus-Peter
KP
Delbrück
Heinrich
H
Behlke
Joachim
J
Müller
Eva-Christina
EC
Jarosch
Ernst
E
Sommer
Thomas
T
Heinemann
Udo
U
D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D019281:Dimerization; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D019308:Palmitic Acid / Q000737:chemistry; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity; D033921:Vesicular Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
C098389:BET3 protein, S cerevisiae; D008565:Membrane Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; C406327:transport protein particle, TRAPP; D019308:Palmitic Acid