Dataset Information

Structural basis for the Pr-Pfr long-range signaling mechanism of a full-length bacterial phytochrome at the atomic level

ABSTRACT: Light sensing mechanisms allow prokaryotes and eukaryotes to adapt to constantly changing environmental factors. Phytochromes constitute a widespread biological photoreceptor family that typically interconvert between two photostates called Pr (red light-absorbing) and Pfr (far-red light-absorbing). Despite the vast structural information reported on phytochromes, the lack of full-length structures solved at the (near-)atomic level in both pure Pr and Pfr states leaves gaps in the structural mechanisms involved in the signal transmission pathways during the photoconversion. Here we present the crystallographic structures of three versions from the plant pathogen Xanthomonas campestris virulence regulator XccBphP bacteriophytochrome, including two full-length proteins, in the Pr and Pfr states. The structures show a reorganization of the interaction networks within and around the chromophore-binding pocket, an α-helix/β-sheet tongue transition, specific domain reorientations, along with interchanging kinks and breaks at the helical spine as a result of the photoswitching, which subsequently affect the quaternary assembly of the protein. These structural findings, combined with mutational, biochemical and computational studies, allow us to describe the signaling mechanism of a full-length bacterial phytochrome at the atomic level. As the transduction mechanism herein reported involves elements highly conserved in other phytochromes, it might be extended to other members of this family of photoreceptors.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Xanthomonas Campestris

SUBMITTER: Germán Rosano

LAB HEAD: Germán Rosano

PROVIDER: PXD026829 | Pride | 2021-12-17

REPOSITORIES: Pride

ACCESS DATA

Similar Datasets

Project description:DM64 is a toxin-neutralizing serum glycoprotein isolated from the South American opossum Didelphis aurita. This ophiophagous marsupial is naturally resistant to snake envenomation and has evolved such biochemical defense as a trophic adaptation. The endogenous antitoxin is a human α1B-glycoprotein homolog of 64 kDa, specifically targeting myotoxic phospholipases A2, which account for most local tissue damage associated with viper snakebites. This study investigated the noncovalent complex formed between native DM64 and myotoxin II, a myotoxic calcium-independent phospholipase-like protein from Bothrops asper venom. Analytical ultracentrifugation, size exclusion chromatography, and small-angle X-ray scattering (SAXS) data indicated that DM64 is monomeric in solution and binds equimolar amounts of the toxin. Initial attempts to solve the structure of DM64 experimentally failed due to technical limitations caused by N-glycan heterogeneity and low recovery of heterologous protein. Classical molecular modeling techniques were impaired by the lack of templates with more than 25% sequence identity with DM64. An integrative structural biology approach was then applied to generate a refined three-dimensional model of the inhibitor bound to myotoxin II. The five immunoglobulin-like domains of DM64 were individually modeled using the I-TASSER suite. Distance constraints generated by cross-linking mass spectrometry of the complex guided the docking of DM64 domains to the crystal structure of myotoxin II (1CLP), using the Rosetta docking suite. The model of the DM64-myotoxin II complex was successfully fitted to a SAXS envelope. Inter-protein cross-links and limited hydrolysis analyses shed light on the inhibitor's regions involved with toxin interaction, revealing the critical participation of the first, third, and fifth domains of DM64. Our data showed that the fifth domain of DM64 binds to myotoxin II’s amino-terminal and -wing regions. The third domain of the inhibitor acts in a complementary way to the fifth domain. Their binding to these toxin regions presumably interferes with dimerization, a critical step in the allosteric activation of myotoxic PLA2-like proteins. Additionally, the first domain of DM64 likely interacts with the functional site of the toxin putatively associated with membrane anchorage. We propose that both mechanisms concur to effectively inhibit myotoxin II toxicity by DM64 binding. In summary, we argue such topological characterization of this toxin-antitoxin complex to constitute an essential first step toward the rational design of novel peptide-based antivenom therapies targeting snake venom myotoxins.

Dataset Information

Structural basis for the Pr-Pfr long-range signaling mechanism of a full-length bacterial phytochrome at the atomic level

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure