High-resolution crystal structure of theBorreliella burgdorferiPlzA protein in complex with c-di-GMP: new insights into the interaction of c-di-GMP with the novel xPilZ domain

Abstract

In the tick-borne pathogens, Borreliella burgdorferi and Borrelia hermsii, c-di-GMP is produced by a single diguanylate cyclase (Rrp1). In these pathogens, the Plz proteins (PlzA, B and C) are the only c-di-GMP receptors identified to date and PlzA is the sole c-di-GMP receptor found in all Borreliella isolates. Bioinformatic analyses suggest that PlzA has a unique PilZN3-PilZ architecture with the relatively uncommon xPilZ domain. Here we present the crystal structure of PlzA in complex with c-di-GMP (1.6 Å resolution). This is the first structure of a xPilz domain in complex with c-di-GMP to be determined. PlzA has a two domain structure, where each domain comprises topologically equivalent PilZ domains with minimal sequence identity but remarkable structural similarity. The c-di-GMP binding site is formed by the linker connecting the two domains. While the structure of apo PlzA could not be determined, previous fluorescence resonance energy transfer data suggest that apo and holo forms of the protein are structurally distinct. The information obtained from this study will facilitate ongoing efforts to identify the molecular mechanisms of PlzA-mediated regulation in ticks and mammals.

Borrelia burgdorferiPlzA is a cyclic-di-GMP dependent DNA and RNA binding protein

“…Without bound c-di-GMP, PlzA has a structure that is flexible, potentially preventing the interactions necessary for DNA binding (Figure 11A left). This hypothesis is supported by the fact that crystallization of PlzA was only achieved with c-di-GMP bound (41). We surmise that binding of c-di-GMP to the C-terminal PilZ domain results in a conformational change of PlzA into a more rigid structure permitting nucleic acid binding through residues of the N-terminal domain (Figure 11A middle and right).…”

“…A common mechanism of action for c-di-GMP effector proteins is to bind nucleic acids and function as transcription factors and/or nucleoid-associated proteins (34)(35)(36)(37)(38)(39)(40). The recently solved crystal structure of B. burgdorferi PlzA revealed a unique dual-domain topology consisting of an amino-terminal PilZ-like domain, called PilZN3, which is connected to the carboxy-terminal PilZ domain via a linker domain that contains the RxxxR c-di-GMP binding motif (41). Binding of c-di-GMP to the interdomain linker domain results in a conformational change;; locking the PlzA domains into a rigid conformation (26,29,41).…”

“…The recently solved crystal structure of B. burgdorferi PlzA revealed a unique dual-domain topology consisting of an amino-terminal PilZ-like domain, called PilZN3, which is connected to the carboxy-terminal PilZ domain via a linker domain that contains the RxxxR c-di-GMP binding motif (41). Binding of c-di-GMP to the interdomain linker domain results in a conformational change;; locking the PlzA domains into a rigid conformation (26,29,41). While no canonical DNA binding domain is evident in the PlzA amino acid sequence, we note that many of the borrelial nucleic acid binding proteins our lab has discovered and characterized possess novel binding motifs (31,(41)(42)(43)(44)(45)(46).…”

BosR and PlzA reciprocally regulate RpoS function to sustain Borrelia burgdorferi in ticks and mammals

“…Binding of c-di-GMP by MrkH induces conformational changes that enable it to bind to DNA and the C-terminal domain of RNAP a subunit (a-CTD) (85). As with MrkH, c-di-GMP binding by PlzA brings together its N-terminal PilZN3 and C-terminal PilZ β-barrels and likely positions three positively charged helices within the PilZN3 domain to create a potential interface for DNA binding (25,63,86).…”

“…Binding of c-di-GMP by MrkH induces conformational changes that enable it to bind to DNA and the C-terminal domain of RNAP α subunit (α-CTD) ( 85 ). As with MrkH, c-di-GMP binding by PlzA brings together its N-terminal PilZN3 and C-terminal PilZ β-barrels and likely positions 3 positively charged helices within the PilZN3 domain to create a potential interface for DNA binding ( 25 , 63 , 86 ). Based on an analysis of PlzA-dependent expression of glpF , the prototypical c-di-GMP-regulated, tick-phase gene, Zhang et al ( 87 ) proposed that PlzA interacts directly with RNAP-RpoD.…”