Cyclic dimeric guanosine monophosphate (cyclic-di-GMP) is a ubiquitous bacterial second messenger that regulates several cellular processes, including biofilm formation. Globin-coupled sensor (GCS) proteins contain a diguanylate cyclase (GG(D/E)EF), cyclic-di-GMP synthesizing domain connected to a globin sensor domain via a middle domain of varying length. While a handful of GCS proteins have been characterized, those associated with human pathogens such as Klebsiella variicola have yet to be studied. Herein, the preliminary characterization of a GCS protein from K. variicola (KvGCS) is reported. Stopped-flow O2 dissociation was biphasic, which is consistent with that of other GCS proteins. Cyclic-di-GMP kinetic assays were able to determine a kcat and KM for both the Fe(II)-O2 and Fe(II) states. Using biological small-angle X-ray scattering, a low-resolution structure of the dimer and tetramer conformations of KvGCS was determined. Overall, this work provides information about the full-length GCS protein scaffolding, and insight into the regulation of KvGCS in the presence and absence of oxygen.
