Poster abstracts
Poster number 9 submitted by Qurat Ashraf
Towards elucidating the structure of the Membrane-Adjacent Domain of Cadherin-23, a protein essential for hearing
Qurat Ashraf (The Ohio State University)
Abstract:
Hair cells of the inner ear transform mechanical stimuli into electrical signals that are transmitted to the central nervous system for processing in hearing and balance. This process of mechanotransduction happens when hair-cell actin-filled apical protrusions, called stereocilia, move upon mechanical stimulation and stretch fine protein filaments connecting the tip of stereocilia to their taller neighbors. As these filaments, called tip links, are stretched, they pull and open ion channels to elicit an ionic current that initiates sensory perception. Tip-link filaments are made of homodimers of cadherin-23 (CDH23) and protocadherin-15 (PCDH15), two proteins involved in inherited deafness. CDH23 forms the upper two-thirds of the tip-links and interacts with PCDH15 at the lower end to form a hetero-tetrameric structure. CDH23 and PCDH15, calcium-dependent cell-cell adhesion proteins, have multiple extracellular cadherin (EC) “repeats”, each about 100 amino acids long with a characteristic Greek-key fold. In addition to these EC repeats, both CDH23 and PCDH15 have membrane-adjacent domains, MAD28 and MAD12, respectively, which are predicted to be similar in structure. Intriguingly, biochemical assays and crystal structures of PCDH15 reveal that MAD12 induces parallel dimerization and that it has a ferredoxin-like fold, unlike the Greek key fold for EC repeats. Yet the structure and biochemical properties of MAD28 and its adjacent CDH23 EC repeats are unknown. To determine whether CDH23 MAD28 has a ferredoxin-like fold that facilitates parallel dimerization, we tested several protocols to express and purify various CDH23 fragments that include EC25 to MAD28. We are now able to express and purify milligram amounts of well-refolded protein amenable for ongoing studies on the oligomerization state of CDH23 and the structure of MAD28. Results from these experiments will advance our understanding of the role played by CDH23 in inner-ear mechanotransduction.
References:
Jaiganesh, A. et al., (2018). Zooming in on cadherin-23: Structural diversity and potential mechanisms of inherited deafness. Structure, 25:1210-1225.
De-la-Torre, P. et al., (2018). A Mechanically Weak Extracellular Membrane-Adjacent Domain Induces Dimerization of Protocadherin-15. Biophysics Journal, 115: 2368-2385.
Choudhary, D. et al., (2020). Structural Determinants of Protocadherin-15 Mechanics and Function in Hearing and Balance Perception. PNAS.
Keywords: structural biology, X-ray crystallography, Biochemistry