Structural study of X-ray induced activation of carbonic anhydrase.

Carbonic anhydrase, a zinc metalloenzyme, catalyzes the reversible hydration of carbon dioxide to bicarbonate. It is involved in processes connected with acid-base homeostasis, respiration, and photosynthesis. More than 100 distinct human carbonic anhydrase II (HCAII) 3D structures have been generated in last 3 decades [Liljas A, et al. (1972) Nat New Biol 235:131-137], but a structure of an HCAII in complex with CO(2) or HCO(3)(-) has remained elusive. Here, we report previously undescribed structures of HCAII:CO(2) and HCAII:HCO(3)(-) complexes, together with a 3D molecular film of the enzymatic reaction observed successively in the same crystal after extended exposure to X-ray. We demonstrate that the unexpected enzyme activation was caused in an X-ray dose-dependent manner. Although X-ray damage to macromolecular samples has long been recognized [Ravelli RB, Garman EF (2006) Curr Opin Struct Biol 16:624-629], the detailed structural analysis reports on X-ray-driven reactions have been very rare in literature to date. Here, we report on enzyme activation and the associated chemical reaction in a crystal at 100 K. We propose mechanisms based on water photoradiolysis and/or electron radiolysis as the main cause of enzyme activation.