Identification of novel peptidyl serine α-galactosyltransferase gene family in plants

Fumie Saito, Akiko Suyama, Takuji Oka, Takehiko Yoko-O, Ken Matsuoka, Yoshifumi Jigami, Yoh Ichi Shimma

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Abstract

In plants, serine residues in extensin, a cell wall protein, are glycosylated withO-linked galactose. However, the enzyme that is involved in the galactosylation of serine had not yet been identified. To identify the peptidyl serine O-α-galactosyltransferase (SGT), we chose Chlamydomonas reinhardtii as a model. We established an assay system for SGT activity using C. reinhardtii and Arabidopsis thaliana cell extracts. SGT protein was partially purified from cell extracts of C. reinhardtii and analyzed by tandem mass spectrometry to determine its amino acid sequence. The sequence matched the open reading frame XP-001696927 in the C. reinhardtii proteome database, and a corresponding DNA fragment encoding 748 amino acids (BAL63043) was cloned from a C. reinhardtii cDNA library. The 748-amino acid protein (CrSGT1) was produced using a yeast expression system, and the SGT activity was examined. Hydroxylation of proline residues adjacent to a serine in acceptor peptides was required for SGT activity. Genes for proteins containing conserved domains were found in various plant genomes, including A. thaliana and Nicotiana tabacum. The AtSGT1 and NtSGT1 proteins also showed SGT activity when expressed in yeast. In addition, knock-out lines of AtSGT1 and knockdown lines of NtSGT1 showed no or reduced SGT activity. The SGT1 sequence, which contains a conserved DXD motif and a C-terminal membrane spanning region, is the first example of a glycosyltransferase with type I membrane protein topology, and it showed no homology with known glycosyltransferases, indicating that SGT1 belongs to a novel glycosyltransferase gene family existing only in the plant kingdom.

Original languageEnglish
Pages (from-to)20405-20420
Number of pages16
JournalJournal of Biological Chemistry
Volume289
Issue number30
DOIs
Publication statusPublished - Jul 25 2014

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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