The βγ-crystallin superfamily contains a Universal motif for binding calcium

Abstract

The βγ-crystallin superfamily consists of evolutionarily related proteins with domain topology similar to lens β- and γ-crystallins, formed from duplicated Greek key motifs. Ca²⁺ binding was found in a few βγ-crystallin members earlier, although its prevalence and diversity as inherent molecular properties among members of the superfamily are not well studied. To increase our understanding of Ca²⁺ binding in various βγ-crystallins, we undertook comprehensive structural and Ca²⁺-binding studies of seven members of the superfamily from bacteria, archaea, and vertebrates, including determination of high-resolution crystal structures of three proteins. Our structural observations show that the determinants of Ca²⁺ coordination remain conserved in the form of an N/D-N/D-#-I-S/T-S motif in all domains. However, binding of Ca²⁺ elicits varied physicochemical responses, ranging from passive sequestration to active stabilization. The motif in this superfamily is modified in some members like lens crystallins where Ca²⁺-binding abilities are partly or completely compromised. We show that reduction or loss of Ca²⁺ binding in members of the superfamily, particularly in vertebrates, is due to the selective presence of unfavorable amino acids (largely Arg) at key Ca²⁺-ligation positions and that engineering of the canonical Ca²⁺-binding residues can confer binding activity on an otherwise inactive domain. Through this work, we demonstrate that βγ-crystallins with the N/D-N/D-#-I-S/T-S motif form an extensive set of Ca²⁺-binding proteins prevalent in all of the three kingdoms of life.