This is a new project initiated during the last quarter of the year. It is an open-ended project that has the potential to develop into a substantial effort. The Zona Pellucida (ZP) is a 7um thick membrane encapsulating the mammalian oocyte. It is known that the ZP is composed of only three glycoproteins (ZP1, ZP2 and ZP3) and that it plays a critical role in fertilization. Neither the structure of any of the three proteins nor the architecture of the ZP microstructure are known although it is believed that ZP1 cross-links the other two proteins, and that ZP2 and ZP3 interact with the sperm during fertilization. It is believed that these interactions controls species-specific fertilization and prevent multiple sperms from reaching the oocyte plasma membrane. ? ? We are using the AFM to image the constituent proteins of solubilized ZPs and investigate the capacity of these proteins to form oligomeric and/or polymeric structures under various physiological conditions. A variety of fiber formations have been observed and the identification of the constituent components of these fibers is being sought using immunochemistry in conjunction with the high resolution imaging capabilities of the atomic force microscope. In addition we are imaging the topography of the outer and inner wall surfaces of the ZP before and after fertilization. Fragments of homogenates allow us to also image inner architecture of the wall structure. Preliminary results indicate that the ZP is a very dilute polymeric network with unique mechanical properties. Viscoelastic moduli of the composite polymeric material are estimated by mathematically analyzing ZP indentation data. Mathematical modeling of a thick elastic spherical shell will also result in an estimate of the mechanical properties of the complete ZP and the associated changes induced by fertilization.
