The present invention relates to the field of genetic engineering, and provides a zipper fastener structure of promoting formation of a protein dimer and application thereof. The zipper fastener can be applied to dimerization of proteins of the same type and dimerization of proteins of different types, and can also be applied to polypeptide cycle formation, polypeptide dimerization, and polypeptide extension. A ESAT6-CFP 10 dimer having an approximately native conformation can be obtained, and the dimer has better solubility, and has a better stimulating effect on memory T cells than a ESAT6-CFP10 fusion protein capable of linear fusion expression. A dimer zipper fastener can assist the formation of a more stable cyclic polypeptide, and a CCP polypeptide added with a dimer fastener can improve the detection rate for citrullinated autoantibodies in serum of a rheumatoid arthritis patient.

Provided herein are methods and markers for diagnosing cardiovascular disease and/or neurodegenearative diseases in a subject. The methods include obtaining a biological sample from a subject in need of diagnosis and detecting the amount of a citrullinated protein or a citrullinated peptide in the biological sample obtained from said subject.

Anti-tumour immune responses to modified self-epitopes. The present invention relates to the use of tumour-associated epitopes in medicine and in particular in the treatment of cancer. The epitopes stimulate an immune reaction against the tumour and have a modification selected from determination of arginine to citrulline, nitration of tyrosine, oxidation of tryptophan and deamination of glutamine or asparagine. The invention also relates to nucleic acids comprising sequences that encode such epitopes for use in the treatment of cancer.

A kit for measuring an anti-cyclic citrullinated peptide antibody, an application thereof, and a test method is provided. The kit includes a biotinylated antigen formed by coupling a plurality of branched peptide units having branched peptide chains using lysine, and the branched peptide units each is formed by coupling a plurality of citrullinated peptides using lysine. The kit and method have high test sensitivity and good specificity; compared with a mixed antigen, citrulline being coupled and then used as a single-component antigen, reduces the complexity of components of the antigen and the difficulty of quality control of the reagent, and improves inter-batch consistency, and quantitative test can be performed and the time needed to complete all procedures to obtain a result is 45 min.

The present invention discloses mass spectrometry (MS)-based methods and tagging reagents for qualitative and quantitative analysis of post-translational modified (PTM) proteins in biological and clinical samples. The present invention utilizes thiol-containing tagging reagents which are able to bind to or derivatize biomolecules, preferably post-translational modified (PTM) polypeptides such as citrullinated and homocitrullinated polypeptides. The tagging reagents are preferably biotin-derived tags able to react with a side chain of the modified polypeptides.

The present disclosure provides methods of selectively label an amino acid residue on a peptide by replacing a post translational modification with a labeling moiety and sequencing the peptide to obtain the location of the amino acid residue and the identity of the post translational modification. In some aspects, the disclosure also provides methods of identifying the position, quantity, the identity of a post translational modification, or any combination thereof, in peptides which may be used for therapeutic purposes.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

Compositions and methods for detection of anti-citrullinated protein antibodies (ACPAs) in rheumatoid arthritis (RA) patients. Patient samples known or suspected of containing ACPAs were probed against citrullinated proteins, and antibody responses to 190 citrullinated proteins in 20 RA patients were investigated. Unique antibody reactivity patterns in both clinical anti-cyclic citrullinated peptide assay positive (CCP+) and negative (CCP) RA patients were observed. At individual antigen levels, six novel antibody/antigen complexes were discovered and validated against specific citrullinated antigens (Myelin Basic Protein (MBP), osteopontin (SPP1), flap endonuclease (FEN1), insulin like growth factor binding protein 6 (IGFBP6), insulin like growth factor I (IGF1) and stanniocalcin-2 (STC2)) in RA patients. Identification of immune-dominant epitope(s) for citrullinated MBP was also performed. The identified biomarkers have high specificity, especially MBP.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

There is provided agents for modulation of a chronic inflammatory response wherein the agent modulates the biological activity of citrullinated tenascin-C. There is also provided methods of identifying agents modulating citrullinated tenascin-C and chronic inflammation. There are also provided therapeutic uses of such agents.