This request for a Shared Instrumentation Grant seeks funds for purchase of a circular dichroism spectropoloarimeter with data processing and computer control capabilities. Use of this instrument will be shared by nine NIH grantees. It will make available the capacity for examination of secondary structure of a variety of proteins and for estimation of the changes in protein conformation upon binding or substitution of metal ions or upon alteration of amino acid sequence by in vitro mutagenesis. The ion-binding proteins include snake venom proteases (Fox), calmodulin (Kretsinger), and several proteins implicated in exocytosis and vesicle fusion (Creutz). Proteins to be examined after mutational alteration include staphylococcal nuclease (Benjamin), the E. coli vitamin B12 receptor (Kadner), histocompatibility antigens (Engelhard), the src oncogene product (Parsons), yeast histones (Smith), and the vesicular stomatits virus integral and peripheral membrane proteins (Pal and Wagner). Information from these studies will be invaluable in showing whether single amino acid substitutions influence the activity of that protein through direct action at the altered site or by disruption or labilization of the structure of the protein or a subdomain. Thus, the requested instrumentation will play an important role in a large number of topically unrelated research projects, none of which would otherwise justify purchase of such equipment. We hope to provide evidence showing the value of this instrumentation in the characterization of genetically engineered proteins.
