Two Distantly Spaced Basic Patches in the Flexible Domain of Huntingtin-Interacting Protein 1 (HIP1) Are Essential for the Binding of Clathrin Light Chain

摘要

The interaction between HIP family proteins (HIP! and HIP12/1R) and clathrin is fundamental to endocytosis. We used circular dichroism (CD) to study the stability of an HIP1 subfragment (aa468-530) that is splayed open. CD thermal melts show HIPI 468-530 is only stable at low temperatures, but this HIPI fragment contains a structural unit that does not melt out even at 83℃. We then created HIP1 mutants to probe our hypothesis that a short hydrophobic path in the opened region is the binding site for clathrin light chain. We found that the binding of hub/LCb was sensitive to mutating two distantly separated basic residues (K474 and K494). The basic patches marked by K474 and K494 are conserved in HIP12/1R. The lack of conservation in sla2p (S. cerevisiae), HIP1 from D. melanogaster, and HIPI homolog ZK370.3 from C. elegans implies the binding of HIP1 and HIP1 homologs to clathrin light chain maybe different in these organisms.