## 3 Results and Discussion
### BLUF Modules in Association with the Effector Domains
#### 3.3.1 EAL and GGDEF Domain  
The EAL and GGDEF domain-containing proteins are widely distributed among bacteria and are involved in the regulation of the cellular level of the universal signaling molecule bis-(3',5')-cyclic-guanosine monophosphate (c-di-GMP), where the former act as diguanylate cyclases (DGCs) while the latter ones are phosphodiesterase (PDEs) [64]. The c-di-GMP generally controls a variety of signaling pathways associated with cell differentiation, bacterial adhesion and biofilm formation, bacterial motility, the colonization of host tissues and virulence [65; 66]. In the EAL-GGDEF domain containing proteins, the GGDEF and EAL motifs in the active sites are crucial for the DGC and PDE enzyme activities [67; 68; 69]. Although most of the proteins involved in c-di-GMP signaling contain the GGDEF/EAL domains as a single polypeptide, possessing both the DGC and PDE enzyme activities, in some cases the GGDEF and EAL domain is also found alone [68]. In some of the EAL-GGDEF domains containing proteins, it was also reported that one [70; 71] or both [72; 73] of these domains are catalytically inactive due to a lack of respective GGDEF and EAL motifs that are crucial for the enzymatic function. In these proteins, the inactive domains either act as regulators [74] or c-di-GMP effectors [72; 73]. [PERSON] and coworker have reported a FimX-like protein (Flip) with a degenerate EAL-GGDEF domain which interacts with the PilZ-Domain protein to control virulence in _Xanthomonas oryzae pv. oryzae_[75]. YhdA is another protein with a degenerate EAL-GGDEF domain that promotes the turnover of CsrB and CsrC (small RNAs), which reduce the expression of the flhDC (flagellar master regulator) by sequestering the CsrA (RNA-binding protein) [76]. Several EAL-GGDEF domains containing proteins are also reported to have N-terminal sensory domains that can regulate the GGDEF and/or EAL domain functions. In the present study, we have selected an EAL only and EAL-GGDEF domain-containing proteins with the blue light using flavin (BLUF) domain as a sensory domain from _E. coli_ and _Thiocystis violascence_, respectively (Table 1). [PERSON] and coworker have characterized the _Klebsiella pneumoniae_ BlrP1 photoreceptor protein biochemically, structurally, mechanistically, and have elucidated the mechanism of the light-induced regulation of the EAL domain (PDE) via the BLUF sensor domain [4]. Upon light illumination, the structural change in the flavin binding pocket (Trp replace Met) of the BLUF domain cross-activates the EAL domain via allosteric communication and increases the PDE activity [4]. The photo-dependent alteration of the BLUF-EAL interactions influences the quaternary structure, the EAL-EAL interface at the dimerization helix, the compound helix \(\alpha\)5 EAL and the loop connecting it to \(\beta\)5 EAL, which are implicated in the EAL activation [4; 77]. In the BlrP1 photoreceptor, the BLUF domain shares a similar architecture, as shown by other BLUF domains from different organisms, where the central BLUF domain (N-terminus) is surrounded by two helices (helical cap; C-terminus) [78; 79; 80]. However, an additional EAL output domain has been identified in the BlrP1 protein, which is connected to the BLUF domain via a 50 A long linker peptide (triosephosphate isomerase (TIM)-barrel fold) [4]. The EAL active site in the BlrP1 photoreceptor involved Glu188, Asn239, Glu272, Asp302, Asp303, Lys323 and Glu359 [4]. We used this well-characterized BlrP1 as a template and aligned it pairwise with EAL only (_E. coli_), EAL-GGDEF (_Thiocystis violascence_ DSM 198), and EAL-PRK15043 domains (_Trichuris trichturis_), and observed that most of the amino acids constituting the EAL active site in BlrP1 are highly conserved through all of the EAL domain-containing proteins (Figure S1), which suggested that the EAL domain-containing proteins possess a similar mechanism for the PDE activity.