Identification Of An Actin Fold Containing Metabolic Enzyme That Undergoes Regulated Polymerization

Abstract

Cytoskeletal actins are thought to have evolved from actin fold enzymes, such as hexokinase or hsp70, that acquired the ability to polymerize while losing all other enzymatic functions apart from ATP hydrolysis. However, the discovery of metabolic enzymes, such as CTP synthetase, whose enzyme activity is regulated by self‐assembly into filaments has suggested another route to evolving cytoskeletal polymers: a filament forming metabolic enzyme that lost its role in metabolism and acquired novel cellular functions. In order to investigate this possibility, we surveyed all of the actin fold sugar kinases in yeast for their ability to form filaments. This screen identified one metabolic actin, the yeast glucokinase, Glk1p, that was capable of forming filaments in vivo. We have reconstituted Glk1p polymerization in vitro and have found that either ATP/Glucose or glucose‐6‐phosphate is sufficient to trigger filament formation. Consistent with this, we have also found that increasing glucose‐6‐phosphate levels in vivo triggers Glk1p polymerization. Furthermore, the addition of glucose and nonhydrolyzable ATP analogs to Glk1p also causes filaments to form. Thus, ATP hydrolysis is not required for Glk1p polymerization. Interestingly, the addition of ADP causes rapid disassembly of glucose‐6‐phosphate/Glk1p filaments suggesting that the stability of the filament is sensitive to the hydrolysis of ATP. While this is reminiscent of the dynamic behavior of cytoskeletal actins, our studies of Glk1p argue that it forms filaments that are structurally distinct from cytoskeletal actin filaments. Since the early steps of glycolysis consume ATP before generating ATP, we propose that Glk1p polymerization acts as a brake on glycolysis to prevent catastrophic depletion of ATP under certain metabolic conditions. We are currently using our structural and biochemical insights into Glk1p polymerization to test this possibility. Because Glk1p is the first filament forming actin that has more enzymatic functions than simple ATP hydrolysis, our results provide novel insights into both the evolution of the actin cytoskeleton and how the plasticity of the actin fold can support both polymerization and essential enzymatic activities.Support or Funding InformationThis work was supported by a HHMI Collaborative Innovation Award