Abstract
The interaction of Tudor domain-containing proteins (TDRDs) with P-element-induced wimpy testis (PIWI) proteins plays critical roles in transposon silencing and spermatogenesis. Most human TDRDs recognize PIWI proteins in a methylarginine-dependent manner via their extended Tudor (eTudor) domains, except TDRD2, which prefers an unmethylated PIWI protein. In order to illustrate the recognition of unmethylated PIWI proteins by TDRD2, we extensively tried co-crystallization of the TDRD2 eTudor with different PIWIL1 peptides, but to no avail. Recombinant antigen-binding fragments (Fabs) have been used to crystallize some difficult proteins in the past, so we generated Fab against the TDRD2 eTudor protein using a phage-display antibody library, and one of these Fab fragments indeed facilitated the co-crystallization of TDRD2 and PIWIL1. Structural analysis of Fab, the TDRD2 eTudor domain in complex with an unmethylated PIWIL1-derived peptide revealed that the PIWIL1 residues G3 through R8 bound between the Tudor core and SN domain of TDRD2. The C-terminal residues of the PIWIL1 peptide were not resolved, presumably due to steric competition with the heavy chain of the Fab. We propose Fab-assisted crystallization as a tool not only for structural studies of single proteins, but also for analysis of interactions between proteins and their ligands in cases where co-crystallization of native protein complexes fails.
Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
