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- Title
Peptide Sequence Mapping around Bisecting GlcNAc-Bearing N -Glycans in Mouse Brain.
- Authors
Ohkawa, Yuki; Kizuka, Yasuhiko; Takata, Misaki; Nakano, Miyako; Ito, Emi; Mishra, Sushil K.; Akatsuka, Haruna; Harada, Yoichiro; Taniguchi, Naoyuki
- Abstract
N-glycosylation is essential for many biological processes in mammals. A variety of N-glycan structures exist, of which, the formation of bisecting N-acetylglucosamine (GlcNAc) is catalyzed by N-acetylglucosaminyltransferase-III (GnT-III, encoded by the Mgat3 gene). We previously identified various bisecting GlcNAc-modified proteins involved in Alzheimer's disease and cancer. However, the mechanisms by which GnT-III acts on the target proteins are unknown. Here, we performed comparative glycoproteomic analyses using brain membranes of wild type (WT) and Mgat3-deficient mice. Target glycoproteins of GnT-III were enriched with E4-phytohemagglutinin (PHA) lectin, which recognizes bisecting GlcNAc, and analyzed by liquid chromatograph-mass spectrometry. We identified 32 N-glycosylation sites (Asn-Xaa-Ser/Thr, Xaa ≠ Pro) that were modified with bisecting GlcNAc. Sequence alignment of identified N-glycosylation sites that displayed bisecting GlcNAc suggested that GnT-III does not recognize a specific primary amino acid sequence. The molecular modeling of GluA1 as one of the good cell surface substrates for GnT-III in the brain, indicated that GnT-III acts on N-glycosylation sites located in a highly flexible and mobile loop of GluA1. These results suggest that the action of GnT-III is partially affected by the tertiary structure of target proteins, which can accommodate bisecting GlcNAc that generates a bulky flipped-back conformation of the modified glycans.
- Subjects
AMINO acid sequence; PEPTIDE mass fingerprinting; PROTEIN structure; SEQUENCE alignment; TERTIARY structure
- Publication
International Journal of Molecular Sciences, 2021, Vol 22, Issue 16, p8579
- ISSN
1661-6596
- Publication type
Article
- DOI
10.3390/ijms22168579