We are at a time in immunology when one wants to understand the detailed chemistry of antibody-antigen union. We are also at a time where we can now tackle this goal by generating monoclonal antibodies against specific antigens and solving the three-dimensional structures of antibody-antigen complexes by x-ray crystallography. To date, there have only been a few antibody crystal structures which have attempted to provide such details. Until recently, these structures have focussed on binding of small haptens to Fab's of multiple myeloma antibodies with unknown specificity. Our present ability to target high affinity antibodies against known antigens enables us to finally study true antibody-antigen interactions. In particular, one wants to understand the structure diversity of antibody recognition of both rigid and flexible antigen structures. This proposal is to determine the three-dimensional structure of high-affinity anti-progesterone antibody Fab' with and without bond steroids. The goal is to understand the molecular interactions which determine the differentiation among antibody binding of progesterone and other steroid hormones. A possible bonus of this work may advance our understanding of the interactions of proteins with steroid hormones. A possible bonus of this work may advance our understanding of the interactions of proteins with steroid hormones. The main objectives of this study are: 1) Crystal Structure of an Anti-progesterone FAB': The three-dimensional structure will be determined from the complete native data set already collected and combined with derivative data to be collected from the presently available high quality Fab crystals. 2) Crystal Structure of an Fab'-Progesterone Complex: Crystals of an antibody-antigen complex have been obtained from a solution of Fab' in the presence of progesterone solubilized by the addition of 5% ethanol. These crystals have the same cell dimensions as the native Fab, diffract to the same resolution but have slightly different morphology and crystallization conditions. 3) Crystal Structure of Fab'-Steroid Complexes. Soaking of preformed Fab crystals in different steroid solutions, or preferably, co-crystallization of the high-affinity steroids with anti-progesterone Fab will be analysed to determine the molecular interactions which contribute to the fine specificity of the steroid hormone-antibody recognition.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Biophysics and Biophysical Chemistry B Study Section (BBCB)
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Scripps Research Institute
San Diego
United States
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