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Site-Directed Conjugation of “Clicked” Glycopolymers To Form Glycoprotein Mimics: Binding to Mammalian Lectin and Induction of Immunological Function

Supporting Info

Jin Geng,^† Giuseppe Mantovani,*^† Lei Tao,^† Julien Nicolas,^† Gaojian Chen,^† Russell Wallis,^§ Daniel A. Mitchell,^‡ Benjamin R. G. Johnson, Stephen D. Evans, and David M. Haddleton*^†

Contribution from the Department of Chemistry, University of Warwick, CV4 7AL, Coventry, United Kingdom; Clinical Sciences Research Institute, Warwick Medical School, University of Warwick, CV2 2DX, Coventry, United Kingdom; Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, United Kingdom; and School of Physics and Astronomy, University of Leeds, Leeds, United Kingdom

J. Am. Chem. Soc., 2007, 129 (49), pp 15156–15163

DOI: 10.1021/ja072999x

Publication Date (Web): November 17, 2007

†

Department of Chemistry, University of Warwick.

In papers with more than one author, the asterisk indicates the name of the author to whom inquiries about the paper should be addressed.

Present address: Laboratoire de Physico-Chimie, Pharmacotechnie et Biopharmacie, UMR CNRS 8612, Univ Paris-Sud, 92296 Chatenay-Malabry Cedex, France.

University of Leicester.

‡

Warwick Medical School, University of Warwick.

University of Leeds.

, D.M.Haddleton@warwick.ac.uk

Abstract

Synthesis of well-defined neoglycopolymer−protein biohybrid materials and a preliminary study focused on their ability of binding mammalian lectins and inducing immunological function is reported. Crucial intermediates for their preparation are well-defined maleimide-terminated neoglycopolymers (M_n = 8−30 kDa; M_w/M_n = 1.20−1.28) presenting multiple copies of mannose epitope units, obtained by combination of transition-metal-mediated living radical polymerization (TMM LRP) and Huisgen [2+3] cycloaddition. Bovine serum albumin (BSA) was employed as single thiol-containing model protein, and the resulting bioconjugates were purified following two independent protocols and characterized by circular dichroism (CD) spectroscopy, SDS PAGE, and SEC HPLC. The versatility of the synthetic strategy presented in this work was demonstrated by preparing a small library of conjugating glycopolymers that only differ from each other for their relative epitope density were prepared by coclicking of appropriate mixtures of mannopyranoside and galactopyranoside azides to the same polyalkyne scaffold intermediate. Surface plasmon resonance binding studies carried out using recombinant rat mannose-binding lectin (MBL) showed clear and dose-dependent MBL binding to glycopolymer-conjugated BSA. In addition, enzyme-linked immunosorbent assay (ELISA) revealed that the neoglycopolymer−protein materials described in this work possess significantly enhanced capacity to activate complement via the lectin pathway when compared with native unmodified BSA.

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