title: Site-specific crosslinking of annexin proteins by 1,4-benzoquinone: a novel crosslinker for the formation of protein dimers and diverse protein conjugates.
abstract: Annexin V (1) specifically binds to phosphatidylserine on apoptotic and necrotic cells as well as certain cancer cells, making it an attractive vehicle for the delivery of therapeutically-relevant conjugates to such sites. The wild-type protein possesses a single thiol at Cys316, which is difficultly accessible to site-specific labeling by simple maleimides. By contrast, 1,4-benzoquinone site-specifically labels annexin V in minutes. The resulting conjugate (5) serves as an intermediate for crosslinking annexin molecules, which can be accomplished within hours either directly for linking annexin V-128 (19), or via an extended sequence involving the crosslinking of two units of (5) by the symmetrical α,ω-dithiol (20). Besides its ability to mediate protein dimer formation while retaining annexin V's ability to bind phosphatidylserine, (5) possesses classic 1,4-benzoquinone reactivity. Various nucleophiles and Diels-Alder dienes form adducts with (5) in reactions that may have general utility for the synthesis of novel biologically active entities. The present work presents the first example of thiol-specific crosslinking of proteins by 1,4-quinone-based methodology designed to exploit the reactivity of this versatile chemical entity.
authors.affiliation: Alcon Research Ltd., 6201 South Freeway, Fort Worth, Texas 76134, USA.
authors.forename: Peng
authors.initials: P
authors.lastname: Yu
authors.forename: Ivona
authors.initials: I
authors.lastname: Strug
authors.forename: Tanya R
authors.initials: TR
authors.lastname: Cafarella
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Seaton
authors.forename: Allen
authors.initials: A
authors.lastname: Krantz
mesh_terms: D017304:Annexin A5 / Q000737:chemistry*; D016227:Benzoquinones / Q000737:chemistry*; D003432:Cross-Linking Reagents / Q000737:chemistry*; D015394:Molecular Structure; D055503:Protein Multimerization
chemical_list: D017304:Annexin A5; D016227:Benzoquinones; D003432:Cross-Linking Reagents; C004532:quinone