Structure of PP4397 Reveals the Molecular Basis for Different c-di-GMP Binding Modes by Pilz Domain Proteins

Ko, Jong Soo; Ryu, Kyoung‐Seok; Kim, Henna; Shin, Jeong Hyun; Lee, Jie‐Oh; Cheong, Chaejoon; Choi, Byong‐Seok

doi:10.1016/j.jmb.2010.03.007

Cited by 94 publications

(122 citation statements)

References 34 publications

(34 reference statements)

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“…Interestingly, the ligand-binding stoichiometry and change in oligomeric state induced upon c-di-GMP binding are distinct from each other, although the same domain organization is employed in both proteins and conserved RxxxR and D/NxSxxG motifs are responsible for c-di-GMP binding. The structure of VCA0042 in complex with c-di-GMP showed that monomeric c-di-GMP was accommodated around the interface between the YcgR-N and PilZ domains [12], whereas binding of two molecules of c-di-GMP induces the dimer-to-monomer transition of PP4397 [13]. Structural analysis of PA4608 from P. aeruginosa, which is one of the single PilZ proteins (PDB: 2L74) [14], revealed that self-intercalated c-di-GMPs observed in the structure of holo-PP4397 were also seen in PA4608.…”

Section: Introductionmentioning

confidence: 99%

The c-di-GMP recognition mechanism of the PilZ domain of bacterial cellulose synthase subunit A

Fujiwara

Komoda

Sakurai

et al. 2013

Biochemical and Biophysical Research Communications

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Section: Introductionmentioning

confidence: 99%

The c-di-GMP recognition mechanism of the PilZ domain of bacterial cellulose synthase subunit A

Fujiwara

Komoda

Sakurai

et al. 2013

Biochemical and Biophysical Research Communications

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“…These cross peaks imply that two c-di-GMP molecules bind to PA4608 with a mutually intercalated conformation similar to that of holo-PP4397. 8 Thus, based on this information and the chemical shift perturbation experiment, 11 a model structure for PA4608 in complex with c-di-GMP was generated (Fig. 4).…”

Section: Two Mutually Intercalated C-di-gmp Molecules Bind To Pa4608mentioning

confidence: 99%

“…Binding of two c-di-GMP molecules induces a dimerto-monomer transition, and the monomeric structure of holo-PP4397 shows that the YcgR-N domain covers the c-di-GMP binding site in the monomeric state. 8 In contrast to apo-PP4397, the dimeric interface of apo-VCA0042 is located in the YcgR-N domain; VCA0042 retains its dimeric state before and after binding of a single c-di-GMP molecule. The binding surface of the VCA0042 PilZ domain is not covered by the YcgR-N domain in the absence of c-di-GMP, whereas it is tightly covered as a result of the inter-domain reorientation between the YcgR-N and PilZ domains in the presence of c-di-GMP.…”

Section: Structure Comparison Among Pilz Domainsmentioning

confidence: 99%

“…7 These differences in binding stoichiometry and oligomeric state are expected to play a role in generating diverse forms of c-di-GMP-mediated regulation. 8 This manuscript describes the molecular characterization of c-di-GMP binding to Pseudomonas aeruginosa PA4608. PA4608 is a single PilZ domain protein whose NMR structure (PDB ID, 1YWU) was determined previously by Ramelot.…”

Section: Introductionmentioning

confidence: 99%

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Structural characterization reveals that a PilZ domain protein undergoes substantial conformational change upon binding to cyclic dimeric guanosine monophosphate

Shin

Ryu

et al. 2010

Protein Science

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PA4608 is a single PilZ domain protein from Pseudomonas aeruginosa that binds to cyclic dimeric guanosine monophosphate (c-di-GMP). Although the monomeric structure of unbound PA4608 has been studied in detail, the molecular details of c-di-GMP binding to this protein are still uncharacterized. Hence, we determined the solution structure of c-di-GMP bound PA4608. We found that PA4608 undergoes conformational changes to expose the c-di-GMP binding site by ejection of the C-terminal 3 10 helix. A dislocation of the C-terminal tail in the presence of c-di-GMP implies that this region acts as a lid that alternately covers and exposes the hydrophobic surface of the binding site. In addition, mutagenesis and NOE data for PA4608 revealed that conserved residues are in contact with the c-di-GMP molecule. The unique structural characteristics of PA4608, including its monomeric state and its ligand binding characteristics, yield insight into its function as a c-di-GMP receptor.

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“…26b). A basic 63 residue in this position is likely necessary to stabilize the interaction with a c-di-GMP dimer, as demonstrated by mutagenesis studies on isolated PilZ domains (Ko et al, 2010;Fujiwara et al, 2013).Most structures of β-barrel-containing PilZ domains contain a short α-helix that follows the last strand of the β−barrel and lays flat across its opening (Benach et al, 2007;Ko et al, 2010). In BcsA, this helix (termed hinge helix) is sandwiched at the interface between the β-barrel and the GT domain (Fig.…”

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confidence: 97%

Structural Studies of Biopolymer Membrane Transport

Morgan¹

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The selective transport of specific substrates across the membrane is an essential part of biology.Organisms have evolved a basic mechanism for the transport of ions and small molecule such as sugars across the membrane. This mechanism is termed 'alternating access' and involves the alternating exposure of a substrate-binding site to either side of the membrane. While the molecular details of 'alternating access' have been revealed for a number of different transporters, this scheme seems infeasible for the transport of long biopolymers such as nucleic acids, proteins, and polysaccharides because 'alternating access' requires that the transporter forms a binding site that is large enough to bind the entire substrate. Here, I present novel data addressing the transport mechanism for two bacterial biopolymer transporter proteins: Bacterial Cellulose Synthase which synthesizes and secretes the polysaccharide, cellulose; and PrtD, an ABC transporter which is involved in the secretion of an extracellular protease.Cellulose is a linear polymer of glucose units that forms a major component of plant cell walls as well as many bacterial biofilms. In bacteria, the components required for cellulose biosynthesis are encoded in a single operon minimally made up of the genes bcsA, bcsB, bcsC, and bcsZ. BcsA is an integral inner-membrane (IM) glycosyltransferase enzyme that is responsible for coupling the synthesis of cellulose with its transport across the IM. BcsB is a periplasmic protein with Cterminal IM anchor, and BcsB forms a complex with BcsA (BcsA-B) that is sufficient for in vitro cellulose synthesis. BcsZ is a periplasmic cellulase enzyme, and BcsC is an outer-membrane β-barrel that presumably forms a pore for the cellulose polymer to cross the outer membrane.iii BcsA-B activity is stimulated by the bacterial signaling molecule, cyclic-di-GMP (c-di-GMP), which is a key regulator of biofilm formation. I present crystal structures of the c-di-GMP-bound BcsA-B complex in the presence and absence of UDP, a competitive inhibitor and substrate mimic. The structures reveal that c-di-GMP releases an auto-inhibited state of the enzyme. A salt bridge stabilizes one of the signature c-di-GMP-binding Arg residues in a position to tether a conserved 'gating loop' in front of the active site. The binding of c-di-GMP releases the tether and allows substrate to access the active site. Additionally, the UDP-bound structure reveals an additional role for the 'gating loop' in coordinating substrate at the active site. Functional experiments confirm the structural interpretation by revealing that disruption of the auto-inhibitory salt bridge by mutagenesis generates a constitutively-active cellulose synthase. The mechanistic insights presented here represent the first examples of how c-di-GMP allosterically modulates enzymatic functions.To address the mechanism by which BcsA-B transports cellulose across the IM, I use in crystallo enzymology with the c-di-GMP-bound BcsA-B crystals. Because crystallized BcsA-B is catalytically a...

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Structure of PP4397 Reveals the Molecular Basis for Different c-di-GMP Binding Modes by Pilz Domain Proteins

Cited by 94 publications

References 34 publications

The c-di-GMP recognition mechanism of the PilZ domain of bacterial cellulose synthase subunit A

The c-di-GMP recognition mechanism of the PilZ domain of bacterial cellulose synthase subunit A

Structural characterization reveals that a PilZ domain protein undergoes substantial conformational change upon binding to cyclic dimeric guanosine monophosphate

Structural Studies of Biopolymer Membrane Transport

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