title: Z-band alternatively spliced PDZ motif protein (ZASP) is the major O-linked β-N-acetylglucosamine-substituted protein in human heart myofibrils.
abstract: We studied O-linked β-N-acetylglucosamine (O-GlcNAc) modification of contractile proteins in human heart using SDS-PAGE and three detection methods: specific enzymatic conjugation of O-GlcNAc with UDP-N-azidoacetylgalactosamine (UDP-GalNAz) that is then linked to a tetramethylrhodamine fluorescent tag and CTD110.6 and RL2 monoclonal antibodies to O-GlcNAc. All three methods showed that O-GlcNAc modification was predominantly in a group of bands ~90 kDa that did not correspond to any of the major myofibrillar proteins. MALDI-MS/MS identified the 90-kDa band as the protein ZASP (Z-band alternatively spliced PDZ motif protein), a minor component of the Z-disc (about 1 per 400 α-actinin) important for myofibrillar development and mechanotransduction. This was confirmed by the co-localization of O-GlcNAc and ZASP in Western blotting and by immunofluorescence microscopy. O-GlcNAcylation of ZASP increased in diseased heart, being 49 ± 5% of all O-GlcNAc in donor, 68 ± 9% in end-stage failing heart, and 76 ± 6% in myectomy muscle samples (donor versus myectomy p < 0.05). ZASP is only 22% of all O-GlcNAcylated proteins in mouse heart myofibrils.
authors.affiliation: Myocardial Function Section, National Heart and Lung Institute, Imperial College London, London W12 0NN, United Kingdom.
authors.forename: Man-Ching
authors.initials: MC
authors.lastname: Leung
authors.forename: Paul G
authors.initials: PG
authors.lastname: Hitchen
authors.forename: Douglas G
authors.initials: DG
authors.lastname: Ward
authors.forename: Andrew E
authors.initials: AE
authors.lastname: Messer
authors.forename: Steven B
authors.initials: SB
authors.lastname: Marston
mesh_terms: D000117:Acetylglucosamine / Q000737:chemistry*; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism / Q000502:physiology*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods; D005786:Gene Expression Regulation; D006321:Heart / Q000502:physiology*; D006801:Humans; D060588:LIM Domain Proteins / Q000378:metabolism / Q000502:physiology*; D008856:Microscopy, Fluorescence / Q000379:methods; D008969:Molecular Sequence Data; D009210:Myofibrils / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D015398:Signal Transduction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000911:Antibodies, Monoclonal; C120696:LDB3 protein, human; D060588:LIM Domain Proteins; D010455:Peptides; D000117:Acetylglucosamine