The protein encoded by the exc-4 locus of C. elegans is an ortholog of the human CLIC (Chloride Intracellular Channel) family of proteins. In exc-4 mutants (excretory canal abnormal), the tubular architecture of the excretory cell lumen is disrupted by cysts. This disease models human tubulocystic kidney disease. Members of the CLIC family are small proteins (~30kDa) that have the unconventional property of translocating from a globular cytosolic form to an integral membrane form, often displaying chloride channel activity. The vertebrate and invertebrate CLIC proteins share both sequence and structural homology with the omega GST family and exhibit a typical N-terminal thioredoxin fold domain. In addition, EXC4 shows several unique features including the absence of the redox sensitive cysteine, an elongated C-terminal extension and a divalent metal binding site. The first 66 amino acid residues are crucial for directing EXC4 to the membrane. The crystal structure of the soluble EXC-4 has been solved. However, due to various limitations of high-resolution structural techniques, the EXC4 membrane structure still remains unsolved. To address these limitations we will use low-resolution biophysical techniques including SDSL-EPR (Site Directed Spin Labeling – Electron Paramagnetic Resonance Spectroscopy), and CD (Circular Dichroism). Cysteine mutants are created by the Site Directed Mutagenesis method to determine the conformation of EXC-4 in the membrane bilayer. We have also generated a C-terminal truncated mutant (EXC4: 1-66) to test how this region mediates the protein insertion into the membrane.