Crystal structure of human carbonic anhydrase‐related protein VIII reveals the basis for catalytic silencing

Abstract

Carbonic anhydrases (CAs; EC 4.2.1.1) catalyze the reversible hydration of carbon dioxide to bicarbonate and a proton.1 They are ubiquitous in prokaryotes and eukaryotes, and are encoded by four evolutionarily-unrelated gene classes (a, b, g, d). The human CA family belongs to the a class and contains 12 catalytic isozymes with different tissue distribution, subcellular localization and kinetic properties. These include the cytosolic (CA I, CA II, CA III, CA VII, CA XIII), membrane-bound (CA IV, CA IX, CA XII, CA XIV), mitochondrial (CA VA, CA VB) and secreted (CA VI) isoforms. They share an absolute requirement for a catalytic Zn ion in the active site, coordinated by an hydroxide ion and three invariant histidine residues H94CAII, H96CAII and H119CAII (human CA II numbering) that are in turn hydrogen bonded to conserved partners Q92CAII, N244CAII and E117CAII, respectively. CAs participate in physiological processes such as respiration, metabolite biosynthesis and pH regulation, and are interesting pharmaceutical targets.2 Sulphonamide-based CA inhibitors are established diuretics and anti-glaucoma drugs, and may have further therapeutic potentials in anti-obesity and anti-cancer treatment.3 The human CA family includes a subclass of three noncatalytic isoforms (CA VIII, CA X, CA XI), also known as CA-related proteins (CA-RPs), based on sequence homology with the catalytic isozymes. CA-RPs lack one or more of the essential Zn-coordinating histidines and are devoid of CO2 hydration activity.4 To date, the biological functions of CA-RPs remain undefined. The first identified CA-RP, CA VIII, replaces the Zn-coordinating H94CAII and its hydrogen-bonding partner Q92CAII with R116 and E114, respectively (human CA VIII numbering).5 CA VIII is highly expressed in the cerebellum,6 and a mouse gene deletion causes a motor coordination defect.7 Relevant to this, CA VIII has been identified as a binding partner for the inositol 1,4,5 triphosphate (IP3) receptor type 1 which is abundant in the cerebellum.8 To provide insights into the biological properties of CA-RPs, we have determined the 1.6 A crystal structure of human CA VIII (hCA VIII). This work represents the first structural characterization of a CA-RP and offers a structural basis for its catalytic silencing effect.