X-ray diffraction studies of two dimeric variants of human pancreatic ribonuclease

Ribonucleases catalyze RNA cleavage reactions. A specialized class of RNases shows a high cytotoxicity toward tumor cell lines, critically dependent on the ability of these molecules to evade the action of the cytosolic ribonuclease inhibitor (RI). Here, we report the X-ray structure of two dimeric variants of RNase1: HHP2-RNase1 and des(16-20)RNase1. HHP2-RNase1 is a covalent dimeric protein, cytotoxic for several malignant cell lines, in which RNase1 has been engineered to reproduce the sequence of bovine seminal ribonuclease helix-II and to eliminate a negative charge on the surface. Des(16-20)RNase1 is a highly stable domain swapped dimer constituted by chains in which five residues in the loop linking the N-terminal helix of RNase1 to the rest of the protein have been deleted. The analysis of HHP2-RNase crystals indicates structural details which can explain the high antitumor activity of the protein. The structure of des(16-20)RNase1 shows a tetrameric association of two swapped dimers, that suggests a pathway of large-scale oligomerization of the protein.