鉴定与硫酸角质素结合的免疫球蛋白互补位并表达用于成像的单链衍生物

Identification of an Immunoglobulin Paratope Binding to Keratan Sulfate and Expression of a Single-Chain Derivative for Imaging.

作者信息

Boyraz Burak, Tauber Rudolf, Dernedde Jens

机构信息

Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin and Berlin Institute of Health, Augustenburger Platz 1, 13353 Berlin, Germany.

Fachbereich Biologie, Chemie, Pharmazie, Freie Universität Berlin, Arnimallee 22, 14195 Berlin, Germany.

出版信息

Biomolecules. 2025 Jan 25;15(2):178. doi: 10.3390/biom15020178.

DOI:10.3390/biom15020178

PMID:40001481

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11852928/

Abstract

Keratan sulfate (KS) is a negatively charged carbohydrate linked to proteins. Several KS-bearing structural glycosaminoglycans participate to maintain the homeostasis of a functional extracellular matrix. Dysfunction of its biochemical composition and structure might therefore lead to pathological situations. For this reason, imaging of KS in tissues is an important diagnostic tool. Here, we describe the identification of the KS paratope derived from the ancestral anti-KS IgG mAb MZ15, as well as the engineering, functional recombinant expression in , and purification of an anti-KS single-chain variable fragment (ScFv). The ScFv enabled in vitro imaging of KS in cryosections of rat cornea by immunofluorescence microscopy comparable to the ancestral IgG MZ15.

摘要

硫酸角质素（KS）是一种与蛋白质相连的带负电荷的碳水化合物。几种含KS的结构糖胺聚糖参与维持功能性细胞外基质的稳态。因此，其生化组成和结构的功能障碍可能会导致病理情况。出于这个原因，组织中KS的成像成为一种重要的诊断工具。在这里，我们描述了源自祖传抗KS IgG单克隆抗体MZ15的KS互补决定区的鉴定，以及抗KS单链可变片段（ScFv）的工程化、在大肠杆菌中的功能性重组表达及纯化。该ScFv通过免疫荧光显微镜能够对大鼠角膜冰冻切片中的KS进行体外成像，效果与祖传IgG MZ15相当。