Abstract
TPST1 is a human tyrosylprotein sulfotransferase that uses 3'phosphoadenosine-5'phosphosulfate (PAPS) to transfer the sulfate moiety to proteins predominantly designated for secretion. To achieve a general understanding of the cellular role of human tyrosine-directed sulfotransferases, we investigated targeting, structure and posttranslational modification of TPST1. Golgi localisation of the enzyme in COS-7 and HeLa cells was visualised by fluorescence imaging techniques. PNGase treatment and mutational studies determined that TPST1 bears N-linked glycosyl residues exclusively at position Asn60 and Asn262. By alanine mutation of these asparagine residues, we could determine that the N-linked oligosaccharides do not have an influence on Golgi retention of TPST1. In concert with N and C-terminal flanking residues, the transmembrane domain of TPST1 was determined to act in targeting and retention of the enzyme to the trans-Golgi compartment. This domain exhibits a pronounced secondary structure in a lipid environment. Further in vivo FRET studies using the transmembrane domain suggest that the human tyrosylprotein sulfotransferase may be functional as homodimer/oligomer in the trans-Golgi compartment.
Original language | English |
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Pages (from-to) | 436-49 |
Number of pages | 14 |
Journal | Journal of Molecular Biology |
Volume | 361 |
Issue number | 3 |
DOIs | |
Publication status | Published - 18 Aug 2006 |
Keywords
- Amino Acid Sequence
- Animals
- Asparagine
- Autoantigens
- COS Cells
- Cercopithecus aethiops
- Dimerization
- Fluorescence Resonance Energy Transfer
- Glycosylation
- Golgi Apparatus
- HeLa Cells
- Humans
- Intracellular Membranes
- Molecular Sequence Data
- Mutation
- Oligosaccharides
- Protein Binding
- Protein Processing, Post-Translational
- Protein Structure, Tertiary
- Sulfotransferases