title: Designing an Antibody-Based Chaperoning System through Programming the Binding and Release of the Folding Intermediate.
abstract: The protein folding pathway consists of sequential intramolecular interactions, while chaperones exert their functions either by stabilizing folding intermediates or by preventing nonspecific intermolecular interactions, which are often associated with aggregation involving exposed hydrophobic residues in folding intermediates. As chaperones do not possess specificity for individual client proteins, we designed an antibody-based chaperoning system to mimic the sequential binding and release of client proteins undergoing folding. The single-chain variable fragment of antibody (scFv) A4 binds to human muscle creatine kinase (HCK) and prevents it from aggregating. The slow dissociation of HCK from A4 resulted in delayed but eventually high-quality refolding, as reflected by the higher recovery of enzymatic activity as well as abolished aggregation. Peptide P6, a sequence in HCK involved in A4 binding, competes with HCK, promotes its dissociation from A4, and accelerates the rate of high-quality refolding. The sequential addition of A4 and P6 is essential for the chaperoning effect. The programmed binding/release method can also be applied to refold HCK from inclusion bodies. Because the association/dissociation of the folding intermediate with the antibody is highly specific, the method can be used to design tailored refolding systems and to investigate chaperoning effects on protein folding/aggregation in a sequence-specific manner.
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Beijing Normal University , Gene engineering and Biotechnology Beijing Key Laboratory, The Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, Beijing 100875, P. R. China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Liu
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Beijing Normal University , Gene engineering and Biotechnology Beijing Key Laboratory, The Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, Beijing 100875, P. R. China.
authors.forename: Caixian
authors.initials: C
authors.lastname: Sun
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Beijing Normal University , Gene engineering and Biotechnology Beijing Key Laboratory, The Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, Beijing 100875, P. R. China.
authors.forename: Cong
authors.initials: C
authors.lastname: Li
authors.affiliation: Korean Bioinformation Center (KOBIC), Korea Research Institute of Bioscience and Biotechnology , Daejeon 305-806, Korea.|Department of Nanobiotechnology and Bioinformatics, University of Sciences and Technology , Daejeon 305-350, Korea.
authors.forename: Jinhyuk
authors.initials: J
authors.lastname: Lee
authors.affiliation: Zhejiang Provincial Key Laboratory of Applied Enzymology, Yangtze Delta Region Institute of Tsinghua University , Jiaxing 314006, P. R. China.
authors.forename: Yong-Doo
authors.initials: YD
authors.lastname: Park
authors.affiliation: B CUBE Center for Molecular Bioengineering, Technische Universität Dresden , Dresden 01307, Germany.
authors.forename: Yixin
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Life Sciences, Beijing Normal University , Gene engineering and Biotechnology Beijing Key Laboratory, The Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, Beijing 100875, P. R. China.
authors.forename: Sen
authors.initials: S
authors.lastname: Li
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry*; D018833:Chaperonins / Q000737:chemistry*; D003402:Creatine Kinase / Q000276:immunology; D006801:Humans; D011485:Protein Binding; D017510:Protein Folding; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology
chemical_list: D000906:Antibodies; D057127:Single-Chain Antibodies; D003402:Creatine Kinase; D018833:Chaperonins