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Impaired O‑Glycosylation at Consecutive Threonine TTX Motifs in Mucins Generates Conformationally Restricted Cancer Neoepitopes
Version 2 2020-03-26, 12:04
Version 1 2020-03-20, 17:42
journal contribution
posted on 2020-03-26, 12:04 authored by Shun Hayakawa, Takahiko Matsushita, Yasuhiro Yokoi, Hajime Wakui, Fayna Garcia-Martin, Hiroshi Hinou, Koji Matsuoka, Kazuhiro Nouso, Toshiya Kamiyama, Akinobu Taketomi, Shin-Ichiro NishimuraAutoantibody signatures
of circulating mucin fragments stem from
cancer tissues, and microenvironments are promising biomarkers for
cancer diagnosis and therapy. This study highlights dynamic epitopes
generated by aberrantly truncated immature O-glycosylation at consecutive
threonine motifs (TTX) found in mucins and intrinsically disordered
proteins (IDPs). NMR analysis of synthetic mucin models having glycosylated
TTX motifs and colonic MUC2 tandem repeats (TRs) containing TTP and
TTL moieties unveils a general principle that O-glycosylation at TTX
motifs generates a highly extended and rigid conformation in IDPs.
We demonstrate that the specific conformation of glycosylated TTX
motifs in MUC2 TRs is rationally rearranged by concerted motions of
multiple dihedral angles and noncovalent interactions between the
carbohydrate and peptide region. Importantly, this canonical conformation
of glycosylated TTX motifs minimizes steric crowding of glycans attached
to threonine residues, in which O-glycans possess
restricted orientations permitting further sugar extension. An antiadhesive
microarray displaying synthetic MUC2 derivatives elicited the presence
of natural autoantibodies to MUC2 with impaired O-glycosylation at
TTX motifs in sera of healthy volunteers and patients diagnosed with
early stage colorectal cancer (CRC). Interestingly, autoantibody levels
in sera of the late stage CRC patients were distinctly lower than
those of early stage CRC and normal individuals, indicating that the
anti-MUC2 humoral response to MUC2 neoepitopes correlates inversely
with the CRC stage of patients. Our results uncovered the structural
basis of the creation of dynamic epitopes by immature O-glycosylation
at TTX motifs in mucins that facilitates the identification of high-potential
targets for cancer diagnosis and therapy.
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Keywords
O-glycosylationMUC 2 neoepitopes correlatesMUC 2 tandemcancer diagnosisTTPTTLMUC 2 derivativesconformationTTX motifsMucins Generates Conformationally Restricted Cancer Neoepitopes Autoantibody signaturesanti-MUC 2 humoral responseIDPmucinstage CRC patientsConsecutive Threonine TTX MotifsNMRglycosylated TTX motifsMUC 2 TRsstage colorectal cancer
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