Truncation of the caspase-related subunit (Gpi8p) of Saccharomyces cerevisiae GPI transamidase: dimerization revealed.
Eukaryotic proteins can be post-translationally modified with a glycosylphosphatidylinositol (GPI) membrane anchor. This modification reaction is catalyzed by GPI transamidase (GPI-T), a multimeric, membrane-bound enzyme. Gpi8p, an essential component of GPI-T, shares low sequence similarity with caspases and contains all or part of the enzyme's active site [U. Meyer, M. Benghezal, I. Imhof, A. Conzelmann, Biochemistry 39 (2000) 3461-3471]. Structural predictions suggest that the soluble portion of Gpi8p is divided into two domains: a caspase-like domain that contains the active site machinery and a second, smaller domain of unknown function. Based on these predictions, we evaluated a soluble truncation of Gpi8p (Gpi8(23-306)). Dimerization was investigated due to the known proclivity of caspases to homodimerize; a Gpi8(23-306) homodimer was detected by native gel and confirmed by mass spectrometry and N-terminal sequencing. Mutations at the putative caspase-like dimerization interface disrupted dimer formation. When combined, these results demonstrate an organizational similarity between Gpi8p and caspases.
Meitzler
Jennifer L
JL
Department of Chemistry, Remsen Hall, Johns Hopkins University, Baltimore, MD 21218, USA.
Gray
Jeffrey J
JJ
Hendrickson
Tamara L
TL
D000595:Amino Acid Sequence; D019881:Aminoacyltransferases; D002240:Carbohydrate Sequence; D020169:Caspases / Q000378:metabolism; D002384:Catalysis; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D019281:Dimerization; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D015815:Cell Adhesion Molecules; D029701:Saccharomyces cerevisiae Proteins; D019881:Aminoacyltransferases; C103133:GPI8 protein, S cerevisiae; D020169:Caspases