Limited proteolysis of native proteins: the interaction between avidin and proteinase K.

摘要: Avidin is a tetramer of 16-kDa subunits that have high affinity for biotin. Proteolysis native apoavidin by proteinase K results in limited attack at the loop between beta-strands 3 and 4, involving amino acids 38-43. Specifically, sites proteolysis are Thr 40-Ser 41 Asn 42-Glu 43. The an avidin product remains otherwise intact which has enhanced binding 4'-hydroxyazobenzene-2-benzoic acid (HABA), chromogenic reporter can occupy biotin-binding site. Saturation site with natural ligand protects from proteolysis, but saturation HABA enhances rate same Analysis three-dimensional structures holoavidin reveals 3-4 accessible to solvent scores highly algorithm developed identify proteolytic attack. structure almost identical apoprotein. In particular, apo holo forms, yet there marked differences susceptibility this region. Evidence suggests rather mobile flexible apoprotein, it becomes constrained upon binding. one crystal appears contacts symmetry-related molecules. Structural analyses suggest "lid" site, formed loop, displaced made more binding, thereby enhancing its susceptibility.