Using selenomethionyl derivatives to assign sequence in low-resolution structures of the AP2 clathrin adaptor

作者：Kelly Bernard T^*; Graham Stephen C; Owen David J

来源：Acta Crystallographica Section D-Structural Biology, 2016, 72(3): 336-345.

DOI：10.1107/S2059798315021580

• selenomethionyl derivatives
• sequence assignment
• low resolution
• AP2 clathrin adaptor complex

Selenomethionine incorporation is a powerful technique for assigning sequence to regions of electron density at low resolution. Genetic introduction of methionine point mutations and the subsequent preparation and crystallization of selenomethionyl derivatives permits unambiguous sequence assignment by enabling the placement of the anomalous scatterers (Se atoms) thus introduced. Here, the use of this approach in the assignment of sequence in a part of the AP2 clathrin adaptor complex that is responsible for clathrin binding is described. AP2 plays a pivotal role in clathrin-mediated endocytosis, a tightly regulated process in which cell-surface transmembrane proteins are internalized from the plasma membrane by incorporation into lipid-enclosed transport vesicles. AP2 binds cargo destined for internalization and recruits clathrin, a large trimeric protein that helps to deform the membrane to produce the transport vesicle. By selenomethionine labelling of point mutants, it was shown that the clathrin-binding site is buried within a deep cleft of the AP2 complex. A membrane-stimulated conformational change in AP2 releases the clathrin-binding site from autoinhibition, thereby linking clathrin recruitment to membrane localization.

• 出版日期2016-3
• 单位河北医科大学