Kvbeta subunits have been shown to affect kinetic properties of voltage-gated K+ channel Kv1alpha subunits and increase the number of cell surface dendrotoxin-binding sites when coexpressed with Kv1. 2. Here, we show that Kvbeta1.2 alters both current expression and gating of Kvalpha1 channels and that each effect is mediated by a distinct Kvbeta1.2 domain. The Kvbeta1.2 N terminus or Kvalpha1-blocking domain introduced steady state current block, an apparent negative shift in steady state activation, and a slowing of deactivation along with a dramatic reduction in single channel open probability. N-terminal deletions of Kvbeta1.2 no longer altered channel kinetics but promoted dramatic increases in Kv1.2 current. The conserved Kvbeta1 C terminus or Kvalpha1 expression domain alone was sufficient to increase the number of functional channels. The same effect was observed with the normally noninactivating subunit, Kvbeta2. By contrast, Kv1.5 currents were reduced when coexpressed with either the Kvbeta1 C terminus or Kvbeta2, indicating that the Kvalpha1 expression domain has Kvalpha1 isoform-specific effects. Our results demonstrate that Kvbeta subunits consist of two domains that are separable on the basis of both primary structure and functional modulation of voltage-gated K+ channels.