Architecture and activation of human muscle phosphorylase kinase

Abstract

The study of phosphorylase kinase (PhK)-regulated glycogen metabolism has contributed to the fundamental understanding of protein phosphorylation. Here we present the high-resolution cryo-electron microscopy structures of human muscle PhK. The 1.3-megadalton PhK 4{beta}4{gamma}4{delta}4 hexadecamer consists of a tetramer of tetramer, wherein four {beta}{gamma}{delta} modules are connected by the central {beta}4 scaffold. The - and {beta}-subunits possess glucoamylase-like domains, but exhibit no detectable enzyme activities. The -subunit serves as a bridge between the {beta}-subunit and the {gamma}{delta} subcomplex, and facilitates the {gamma}-subunit to adopt an autoinhibited state. Ca2+-free calmodulin ({delta}-subunit) binds to the {gamma}-subunit in a compact conformation. Upon binding of Ca2+, a conformational change occurs, allowing for the de-inhibition of the {gamma}-subunit through a spring-loaded mechanism. We also reveal an ADP-binding pocket in the {beta}-subunit, which plays a role in allosterically enhancing PhK activity. These results provide unprecedented molecular insights of this important kinase complex.