Identification and Characterisation of a cyclic di-GMP-specific Diguanylate Cyclase and Phosphodiesterase Genes in Klebsiella Pneumoniae MBB9

Abstract

Bis-(3’–5’)-cyclic-dimeric guanosine monophosphate (c-di-GMP) is a bacterial second messenger known to mediate the regulation of multiple cellular processes, including bacterial adhesion and biofilm formation, bacterial motility, and control the virulence of bacterial pathogens. In many bacteria, the second messenger c-di-GMP, an intracellular signalling molecule, plays a key role in the lifestyle changes and controls the transition between the motile planktonic and sessile biofilm lifestyles. The intracellular levels of c-di-GMP are controlled by c-di-GMP synthesis and degradation catalyzed by diguanylate cyclase (DGC) and phophodiesterase (PDE) enzymes. The GGDEF protein domain synthesizes cyclic di-GMP, whereas the EAL and HD-GYP domains are involved in cyclic di-GMP hydrolysis. Various bacteria contain many copies of these proteins with a diverse organizational structure that highlights the complex regulatory mechanisms of this signaling network. The whole genome of Klebsiella pneumoniae MBB9, recovered from river-stones collected from the Porter Brook, Sheffield, was sequenced and compared to K. pneumoniae 342 to identify DGCs and PDEs and analyze the domain structure of such proteins. Klebsiella pneumoniae MBB9 harboured multiple copies of proteins with GGDEF and EAL domains, most of these were linked to sensory domains and were found to possess 11 genes with GGDEF domains, 11 genes with EAL domains, and 6 genes with both GGDEF and EAL domains. Thirty-nine percent of these proteins contained the GGDEF sequence motif, whereas 39% had EAL sequence motif, and 21 % were hybrid proteins containing both GGDEF and EAL domains. The majority of GGDEF domains are catalytically active as they have an intact conserved A site, whereas all EAL domains have c-di-GMP PDE activity except BluF_2 and YahA proteins.