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Co-Crystallization with Diabodies: A Universal Method for the Introduction of Synthetic Symmetry
21 Pages Posted: 16 Apr 2020 Publication Status: Published
More...Abstract
This work presents a method for introducing synthetic symmetry into protein crystallization samples using an antibody fragment termed a diabody (Dab). These Dabs contain two target binding sites and engineered disulfide bonds have been included to modulate Dab flexibility. The introduction of a disulfide bond promotes a preferred conformation and an increased likelihood of crystallization as observed in solution by small-angle X-ray scattering and in high-resolution crystal structures. To explore the capacity of engineered Dabs as crystallization chaperones, complexes between the engineered Dabs and HIV-1 reverse transcriptase (RT) bound to a high-affinity DNA aptamer were generated. This strategy increased the crystallization hit frequency obtained for RT-aptamer and the structure of a Dab-RT-DNA complex was determined to 3.0 Å resolution. Introduction of synthetic symmetry using a Dab is a broadly applicable strategy, especially when monoclonal antibodies for a target have previously been identified.
Keywords: Diabody, Dab, Co-crystallization, Synthetic Symmetry, Protein Engineering, Crystal Engineering, Reverse Transcriptase
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