Conversion of the enzyme guanylate kinase into a mitotic-spindle orienting protein by a single mutation that inhibits GMP-induced closing

Abstract

New protein functions can require complex sequence changes, but the minimal path is not well understood. The guanylate kinase enzyme (GK enz), which catalyzes phosphotransfer from ATP to GMP, evolved into the GK domain (GK dom), a protein-binding domain found in membrane associate guanylate kinases that function in mitotic spindle orientation and cell adhesion. Using an induced polarity assay for GK dom function, we show that a single serine to proline mutation is sufficient to switch extant GK enz into a functional GK dom. The mutation blocks catalysis (GK enz function) but allows protein binding and spindle orientation (GK dom function). Furthermore, whereas the GK enz undergoes a large closing motion upon GMP binding, fluorescence quenching and NMR demonstrate that the S → P mutation inhibits GMP-induced GK movements. Disrupting GK closing with a mutation at a different position also leads to GK dom function, suggesting that blocking the GK enz closing motion is sufficient for functional conversion of GK enz to GK dom. Although subtle changes in protein function can require complex sequence paths, our work shows that entirely new functions can arise from single mutations that alter protein dynamics.