Abstract
Tristetraprolin (TTP) and TIS11d are two human RNA-binding proteins that belong to the CCCH-type tandem zinc finger family. In the RNA-free state, TIS11d coordinates a zinc ion in each of its two fingers, while TTP coordinates a single zinc ion with the N-terminal zinc finger. We have previously identified three residues, located in the C-terminal half of a short α-helix in the second zinc finger, that control how structured the RNA-binding domain is in these two proteins: Y151, L152, and Q153 in TTP and H201, T202, and I203 in TIS11d. Here, we have used molecular dynamics, NMR spectroscopy, and other biochemical methods to investigate the role of these three residues in the stability of the RNA-binding domain. We found that the intrahelical hydrogen bond formed by the T202 hydroxyl group in the C-terminal zinc finger of TIS11d is necessary to allow for π-π stacking between the side chains of a conserved phenylalanine and the zinc-coordinating histidine. We demonstrated that the lack of this hydrogen bond in TTP is responsible for the reduced zinc affinity of the C-terminal zinc finger.