Specific antibody for identifying K27 ubiquitin chain

By performing structural simulation and site-directed mutagenesis on antibody zl1901, zl1901-R3-S8 was developed, which solves the problem of insufficient specificity of existing antibodies and achieves accurate identification and enrichment of K27 ubiquitin chains, making it suitable for various biological detections and disease diagnosis.

CN122234202APending Publication Date: 2026-06-19XIN HUA HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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Patent Information

Application Number
CN202610311044.9
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2026-03-14
Publication Date
2026-06-19

AI Technical Summary

Technical Problem

Existing technologies lack highly specific antibodies capable of accurately identifying K27 ubiquitin chains. The primary screening antibody zl1901 suffers from insufficient chain-binding specificity and cross-binding with other types of ubiquitin chains, failing to meet the requirements for the identification and enrichment of low-abundance K27-linked ubiquitin chains.

Method used

An antibody, zl1901-R3-S8, was developed through structural simulation and site-directed mutagenesis. The specific mutation sites are T28L, T30F, S31L, Y32R, Y33A, S54I, S57I, R101A, S215V, and S216V. It binds to the CDR3 region of the heavy chain and forms hydrogen bonds with the P19 residue of K27-diUb, thus optimizing it into a full-length IgG form. This improves the affinity for K27-diUb and reduces cross-binding with other types of ubiquitin chains.

Benefits of technology

The antibody zl1901-R3-S8 significantly improves the specific recognition ability of K27 ubiquitin chains, has no cross-binding with other types of ubiquitin chains, exhibits excellent binding affinity, is suitable for a variety of biological assays, has high structural stability, and is applicable to scientific research tools and disease diagnosis.

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Abstract

The invention discloses a specific antibody for identifying a K27 ubiquitin chain, and belongs to the technical field of biological medicines. The antibody is obtained by taking a primary screening antibody zl1901 as a female parent and carrying out structural simulation, molecular docking and site-directed mutagenesis directional transformation, a core optimal strain is zl1901-R3-S8, light chain CDR1 and CDR2 regions of the antibody can be tightly combined with K27-diUb receptor and donor ubiquitin, and a heavy chain CDR3 region and K27-diUb donor ubiquitin P19 residues form a specific hydrogen bond. The affinity KD of the antibody to a K27-connected ubiquitin chain reaches 16.5 nM, the antibody does not have cross binding with M1, K6, K11, K48 and K63-connected ubiquitin chains, and only has extremely weak binding with K29-diUb, so that endogenous and endogenous K27-connected ubiquitination modification of cells can be accurately identified, and a core tool is provided for research on a K27 ubiquitination regulation mechanism.
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