Heme pocket modulates protein conformation and diguanylate cyclase activity of a tetrameric globin coupled sensor.

Bacteria use the second messenger cyclic dimeric guanosine monophosphate (c-di-GMP) to control biofilm formation and other key phenotypes in response to environmental signals. Changes in oxygen levels can alter c-di-GMP signaling through a family of proteins termed globin coupled sensors (GCS) that contain diguanylate cyclase domains. Previous studies have found that GCS diguanylate cyclase activity is controlled by ligand binding to the heme within the globin domain, with oxygen binding resulting in the greatest increase in catalytic activity. Herein, we present evidence that heme-edge residues control O2-dependent signaling in PccGCS, a GCS protein from Pectobacterium carotovorum, by modulating heme distortion. Using enzyme kinetics, resonance Raman spectroscopy, small angle X-ray scattering, and multi-wavelength analytical ultracentrifugation, we have developed an integrated model of the full-length PccGCS tetramer and have identified conformational changes associated with ligand binding, heme conformation, and cyclase activity. Taken together, these studies provide new insights into the mechanism by which O2 binding modulates activity of diguanylate cyclase-containing GCS proteins.