This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We use site directed mutagenesis and cysteine scanning to prepare recombinant peptides for biophysical studies, including spin label EPR, to determine important structural features that contribute toward different affinities of two spectrin isoforms in dimer self-association to form tetramers. The high degrees of sequence homology amongst different isoforms at the tetramerization sites suggest similar self-association affinities. Yet at least two of the better studied isoforms, erythroid and brain spectrins, exhibit quite different affinities. Since several of spectrin?s functions, including coupling of cell adhesion molecules to membranes via a spectrin lattice, as well as maintaining the integrity of erythrocytes, require spectrin to be in the tetramer state, different affinities lead to different tetramer concentrations, and thus to different functions within the biological milieu. The recombinant peptides prepared for these studies are analyzed by mass spectrometry and circular dichroism before biophysical studies to ensure their identities and integ
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