[HTML][HTML] Identification of a Golgi GPI-N-acetylgalactosamine transferase with tandem transmembrane regions in the catalytic domain

T Hirata, SK Mishra, S Nakamura, K Saito… - Nature …, 2018 - nature.com
T Hirata, SK Mishra, S Nakamura, K Saito, D Motooka, Y Takada, N Kanzawa, Y Murakami…
Nature communications, 2018nature.com
Many eukaryotic proteins are anchored to the cell surface via the glycolipid
glycosylphosphatidylinositol (GPI). Mammalian GPIs have a conserved core but exhibit
diverse N-acetylgalactosamine (GalNAc) modifications, which are added via a yet
unresolved process. Here we identify the Golgi-resident GPI-GalNAc transferase PGAP4
and show by mass spectrometry that PGAP4 knockout cells lose GPI-GalNAc structures.
Furthermore, we demonstrate that PGAP4, in contrast to known Golgi glycosyltransferases …
Abstract
Many eukaryotic proteins are anchored to the cell surface via the glycolipid glycosylphosphatidylinositol (GPI). Mammalian GPIs have a conserved core but exhibit diverse N-acetylgalactosamine (GalNAc) modifications, which are added via a yet unresolved process. Here we identify the Golgi-resident GPI-GalNAc transferase PGAP4 and show by mass spectrometry that PGAP4 knockout cells lose GPI-GalNAc structures. Furthermore, we demonstrate that PGAP4, in contrast to known Golgi glycosyltransferases, is not a single-pass membrane protein but contains three transmembrane domains, including a tandem transmembrane domain insertion into its glycosyltransferase-A fold as indicated by comparative modeling. Mutational analysis reveals a catalytic site, a DXD-like motif for UDP-GalNAc donor binding, and several residues potentially involved in acceptor binding. We suggest that a juxtamembrane region of PGAP4 accommodates various GPI-anchored proteins, presenting their acceptor residue toward the catalytic center. In summary, we present insights into the structure of PGAP4 and elucidate the initial step of GPI-GalNAc biosynthesis.
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