The histone variant H2A.Z is a universal mark of gene promoters, enhancers, and 
regulatory elements in eukaryotic chromatin. The chromatin remodeler SWR1 
mediates site-specific incorporation of H2A.Z by a multi-step histone 
replacement reaction, evicting histone H2A-H2B from the canonical nucleosome and 
depositing the H2A.Z-H2B dimer. Binding of both substrates, the canonical 
nucleosome and the H2A.Z-H2B dimer, is essential for activation of SWR1. We 
found that SWR1 primarily recognizes key residues within the α2 helix in the 
histone-fold of nucleosomal histone H2A, a region not previously known to 
influence remodeler activity. Moreover, SWR1 interacts preferentially with 
nucleosomal DNA at superhelix location 2 on the nucleosome face distal to its 
linker-binding site. Our findings provide new molecular insights on recognition 
of the canonical nucleosome by a chromatin remodeler and have implications for 
ATP-driven mechanisms of histone eviction and deposition.