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Engineered synthetic antibodies as probes to quantify the energetic contributions of ligand binding to conformational changes in proteins

Journal Article · · Journal of Biological Chemistry
 [1];  [2];  [2];  [3];  [1];  [4];  [1];  [5]
  1. Univ. of Chicago, IL (United States)
  2. North Illinois Univ., DeKalb, IL (United States)
  3. Indiana Univ., South Bend, IN (United States)
  4. Ridgeview Diagnostics AB, Uppsala (Sweden)
  5. Univ. of Chicago, IL (United States); Gordon Center for Integrative Science, Chicago, IL (United States). Inst. for Biophysical Dynamics
+ Show Author Affiliations
Conformational changes in proteins due to ligand binding are ubiquitous in biological processes and are integral to many biological systems. However, it is often challenging to link ligand-induced conformational changes to a resulting biological function because it is difficult to distinguish between the energetic components associated with ligand binding and those due to structural rearrangements. Consequently, here, we used a unique approach exploiting conformation-specific and regio-specific synthetic antibodies (sABs) to probe the energetic contributions of ligand binding to conformation changes. Using maltose-binding protein (MBP) as a model system, customized phage-display selections were performed to generate sABs that stabilize MBP in different conformational states, modulating ligand-binding affinity in competitive, allosteric, or peristeric manners. We determined that the binding of a closed conformation–specific sAB (sAB-11M) to MBP in the absence of maltose is entropically driven, providing new insight into designing antibody-stabilized protein interactions. Crystal structures of sABs bound to MBP, together with biophysical data, delineate the basis of free energy differences between different conformational states and confirm the use of the sABs as energy probes for dissecting enthalpic and entropic contributions to conformational transitions. Our work provides a foundation for investigating the energetic contributions of distinct conformational dynamics to specific biological outputs. We anticipate that our approach also may be valuable for analyzing the energy landscapes of regulatory proteins controlling biological responses to environmental changes.
Research Organization:
Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Organization:
Chicago Biomedical Consortium; National Institutes of Health (NIH)
OSTI ID:
1432877
Journal Information:
Journal of Biological Chemistry, Journal Name: Journal of Biological Chemistry Journal Issue: 8 Vol. 293; ISSN 0021-9258
Publisher:
American Society for Biochemistry and Molecular BiologyCopyright Statement
Country of Publication:
United States
Language:
ENGLISH

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Cited By (2)

An engineered ultra‐high affinity Fab‐Protein G pair enables a modular antibody platform with multifunctional capability journal November 2019
Structure and development of single domain antibodies as modules for therapeutics and diagnostics journal October 2019

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Related Subjects

59 BASIC BIOLOGICAL SCIENCES
allostery
antibody engineering
conformation-specific synthetic antibodies
crystal structure
energy probes
epitope mapping
phage display
thermodynamics