High reolution structural studies on peripherin, an intergal membrane protein

Abstract

Peripherin is a tetraspannin transmembrane protein localized in retinal rod outer segment disk membranes whose complete 3-dimensional structure is the subject of this nuclear magnetic resonance (NMR) analysis. Peripherin promotes membrane fusion and may be involved in fusion events with rod cell membranes during the rod outer segment disk renewal processes. The 63-residue C-terminal extramembraneous region of peripherin (PERCTER) is crucial to peripherin-mediated fusion and contains both an incisure affinity region and an OS targeting signal. Several mutations in this C-terminal region have been linked to various forms of retinal degenerative disease. The first target of our structural analysis has been the PERCTER, expressed and labeled with 15N, and subjected to structural determination by solution NMR. For this analysis, 3-dimensional HSQC-NOESY and HSQC-TOCSY experiments were utilized, along with 2-dimensional 1H NOESY and TOCSY experiments at various mixing times. This analysis is followed by a similar study of the PERCTER in the presence of a membrane-mimetic, DPC micelles. The next stage of our examination is to obtain NMR data from the isotope-labeled full length peripherin. Detailed understanding of the peripherin structure provides greater insight into retinal degenerative diseases and the mechanisms of membrane fusion. NIH EY10420-14, E. Matilda Ziegler Vision Award.