The mucin-selective protease StcE enables molecular and functional analysis of human cancer-associated mucins.

Autor:	Malaker SA; Department of Chemistry, Stanford University, Stanford, CA 94305., Pedram K; Department of Chemistry, Stanford University, Stanford, CA 94305., Ferracane MJ; Department of Chemistry, University of Redlands, Redlands, CA 92373., Bensing BA; Department of Medicine, San Francisco Veterans Affairs Medical Center and University of California, San Francisco, CA 94143., Krishnan V; Stanford Women's Cancer Center, Division of Gynecologic Oncology, Stanford University, Stanford, CA 94305., Pett C; Leibniz-Institut für Analytische Wissenschaften (ISAS), 44227 Dortmund, Germany.; Department of Chemistry, Umeå University, 901 87 Umeå, Sweden., Yu J; Leibniz-Institut für Analytische Wissenschaften (ISAS), 44227 Dortmund, Germany., Woods EC; Department of Chemistry, Stanford University, Stanford, CA 94305., Kramer JR; Department of Bioengineering, University of Utah, Salt Lake City, UT 84112., Westerlind U; Leibniz-Institut für Analytische Wissenschaften (ISAS), 44227 Dortmund, Germany.; Department of Chemistry, Umeå University, 901 87 Umeå, Sweden., Dorigo O; Stanford Women's Cancer Center, Division of Gynecologic Oncology, Stanford University, Stanford, CA 94305., Bertozzi CR; Department of Chemistry, Stanford University, Stanford, CA 94305; bertozzi@stanford.edu.; Howard Hughes Medical Institute, Stanford, CA 94305.
Jazyk:	angličtina
Zdroj:	Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2019 Apr 09; Vol. 116 (15), pp. 7278-7287. Date of Electronic Publication: 2019 Mar 25.
DOI:	10.1073/pnas.1813020116
Abstrakt:	Mucin domains are densely O -glycosylated modular protein domains that are found in a wide variety of cell surface and secreted proteins. Mucin-domain glycoproteins are known to be key players in a host of human diseases, especially cancer, wherein mucin expression and glycosylation patterns are altered. Mucin biology has been difficult to study at the molecular level, in part, because methods to manipulate and structurally characterize mucin domains are lacking. Here, we demonstrate that secreted protease of C1 esterase inhibitor (StcE), a bacterial protease from Escherichia coli , cleaves mucin domains by recognizing a discrete peptide- and glycan-based motif. We exploited StcE's unique properties to improve sequence coverage, glycosite mapping, and glycoform analysis of recombinant human mucins by mass spectrometry. We also found that StcE digests cancer-associated mucins from cultured cells and from ascites fluid derived from patients with ovarian cancer. Finally, using StcE, we discovered that sialic acid-binding Ig-type lectin-7 (Siglec-7), a glycoimmune checkpoint receptor, selectively binds sialomucins as biological ligands, whereas the related receptor Siglec-9 does not. Mucin-selective proteolysis, as exemplified by StcE, is therefore a powerful tool for the study of mucin domain structure and function. Competing Interests: Conflict of interest statement: A patent application relating to the use of enzymes to digest mucin-domain glycoproteins has been filed by Stanford University (docket no. STAN-1510PRV). C.R.B. is a cofounder and Scientific Advisory Board member of Palleon Pharmaceuticals, Enable Bioscience, Redwood Biosciences (a subsidiary of Catalent), and InterVenn Biosciences, and a member of the Board of Directors of Eli Lilly & Company. O.D. has participated in advisory boards for Tesaro, Merck, and Geneos. O.D. is a speaker for Tesaro and AstraZeneca.
Databáze:	MEDLINE
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