The calgranulins form a class of S100 proteins in higher vertebrates that innate-immune cells release in abundance at infection sites. These proteins function by binding transition metal ions to prevent microbial pathogens from obtaining those essential nutrients. Mammals express three distinct members of this family, S100A8 (calgranulin A), S100A9 (calgranulin B, which heterooligomerizes with S100A8 to form calprotectin), and S100A12 (calgranulin C), that exhibit Ca(II)-dependent transition-metal-binding properties. Human calprotectin effectively sequesters Mn(II), Fe(II), Ni(II), and Zn(II), whereas human S100A12 selectivity sequesters Zn(II) over these other metal ions. Birds and reptiles express a single calgranulin homolog named MRP126, which we reasoned could have properties more similar to either calprotectin or S100A12. Here we present the purification and biophysical characterization of chicken MRP126 and, to the best of our knowledge, provide the first assessment of the metal-binding and antimicrobial properties of an avian MRP126. We show that MRP126 is a homodimer that selectively sequesters Zn(II) and restricts the growth of certain microbes. MRP126 binds Zn(II) at two canonical His(3)Asp sites. The presence of excess Ca(II) increases the affinity of the His(3)Asp sites from the low-nanomolar to the low-picomolar range, thereby enhancing antimicrobial activity. Chicken MRP126 also binds additional Zn(II) equivalents with low-nanomolar affinity at two non-conserved dicysteine sites and with high-nanomolar affinity using a histidine-rich C-terminal tail that is a hallmark of this clade of calgranulins. Our results with chicken MRP126 suggest that Ca(II)-dependent Zn(II) sequestration was a role of the last common ancestor of calgranulin proteins, with mammalian calprotectin subsequently evolving a broader metal-binding repertoire.
