PMID: 8440706Feb 25, 1993Paper

Location of the human red cell spectrin tetramer binding site and detection of a related "closed" hairpin loop dimer using proteolytic footprinting.

The Journal of Biological Chemistry
David W SpeicherL Weglarz

Abstract

Head-to-head association of two spectrin alpha beta heterodimers to form tetramers involves the formation of two equivalent alpha-beta complexes. The sites on the alpha subunit N-terminal region and beta subunit C-terminal region that form these alpha beta complexes have been identified using protease footprinting and direct binding assays. The existence of a similar previously hypothesized internal head-to-head alpha beta interaction in dimers was also demonstrated. The discrete regions of both subunits that are protected from proteolysis in tetramers and dimers are not due to the laterally associated subunit since head-to-head complexes of a univalent alpha peptide with a univalent beta peptide show similar protection of the same sites. These sites are unshielded immediately after monomers assemble side-to-side to form heterodimers, demonstrating that reconstituted dimers are initially in an "open" conformation. Conversion of open dimers to a closed form through formation of the internal head-to-head alpha beta association, as demonstrated by restoration of protease protection, occurred on a time scale of hours at 0 degrees C. Analysis of peptide binding affinities as well as isolation and sequence analysis of head-to-head al...Continue Reading

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