REFINED STRUCTURE OF VILLIN 14T AND A DETAILED COMPARISON WITH OTHER ACTIN-SEVERING DOMAINS

Villin 14T is the amino terminal actin monomer binding domain from the actin-severing and bundling protein villin. Its structure has been de termined in solution using heteronuclear multidimensional nuclear magn etic resonance (NMR) spectroscopy (Markus MA, Nakayama T, Matsudaira P , Wagner G. 1994. Solution structure of villin 14T, a domain conserved among actin-severing proteins. Protein Science 3:70-81). An additiona l nuclear Overhauser effect (NOE) spectroscopy data set, acquired usin g improved gradient techniques, and further detailed analysis of exist ing data sets, produced an additional 601 NOE restraints for structure calculations. The overall fold does not change significantly with the additional NOE restraints but the definition of the structure is impr oved, as judged by smaller deviations among an ensemble of calculated structures that adequately satisfy the NMR restraints. Some of the sid e chains, especially those in the hydrophobic core of the domain, are much more defined. This improvement in the detail of the solution stru cture of villin 14T makes it interesting to compare the structure with the crystal structure of gelsolin segment 1, which shares 58% sequenc e identity with villin 14T, in an effort to gain insight into villin 1 4T's weaker affinity for actin monomers. Villin 14T has smaller side c hains at several positions that make hydrophobic contacts with actin i n the context of gelsolin segment 1. The structure is also compared wi th the structure of the related actin-severing domain, severin domain 2.