Fourier transform infrared linked analysis of conformational changes in annexin V upon membrane binding

Annexins are ubiquitous cellular proteins of unknown primary function that bind to anionic phospholipid membranes in a calcium-dependent manner. Corre lative studies involving X-ray crystallography and electron microscopy sugg est that annexins undergo a structural change upon binding to supported lip id monolayer membranes. in this investigation, novel spectroscopic and anal ytical techniques have been applied to verify and characterize this change. Soluble annexin V was examined with ordinary transmission infrared spectro scopy, while membrane-bound annexin V was examined with both transmission a nd internal reflection infrared spectroscopy. Spectra were processed by lin ked analysis, whereby multiple spectra are fit simultaneously with componen t bands that are constrained to share common fitting parameters. This appro ach is shown to enhance the sensitivity and accuracy of the bandfitting pro cedure. Our results are consistent with the general mode of membrane bindin g inferred from electron microscopy studies, and they provide independent s upport for the conclusion that annexin V undergoes a conformational change upon binding to lipid monolayer membranes. Most likely, this change involve s the formation of new beta structure in which interstrand hydrogen bonds o rient parallel to the membrane surface.